Pyrimidine compounds and uses thereof

ABSTRACT

This invention features pyrimidine compounds of formula (I): 
     
       
         
         
             
             
         
       
     
     R t  is 
     
       
         
         
             
             
         
       
     
     aryl, or heteroaryl; each of R 2  and R 4 , independently, is halogen, nitro, cyano, isothionitro, SR c , or OR c ; or R 2  and R 4 , taken together, is carbonyl; R 3  is R c , alkenyl, alkynyl, OR c , OC(O)R c , SO 2 R c , S(O)R c , S(O 2 )NR c R d , NR c R d , NR c COR d , NR c C(O)OR d , NR c C(O)NR c R d , NR c SO 2 R d , COR c , C(O)OR c , or C(O)NR c R d ; R 5  is H or alkyl; n is 0, 1, 2, 3, 4, 5, or 6; X is O, S, S(O), S(O 2 ), or NR c ; Y is a covalent bond, CH 2 , C(O), C═N—R c , C═N—OR c , C═N—SR c , O, S, S(O), S(O 2 ), or NR c ; Z is N or CH; one of U and V is N, and the other is CR c ; and W is O, S, S(O), S(O 2 ), NR c , or NC(O)R c ; in which each of R a  and R b , independently, is H, alkyl, aryl, heteroaryl; and each of R c  and R d , independently, is H, alkyl, aryl, heteroaryl, cyclyl, heterocyclyl, or alkylcarbonyl. 
     The featured compounds inhibit the production of IL-12, IL-23 and IL-27 and are useful for treating disorders associated with IL-12, IL-23 and IL-27 overproduction or misregulation, such as inflammatory and immune disorders,

RELATED APPLICATIONS

This application claims the benefit of U.S. provisional patentapplication Ser. No. 60/629,505, filed Nov. 19, 2004, the disclosure ofwhich is incorporated herein in its entirety by this reference.

BACKGROUND

Interleukin-12 (IL-12) is a heterodimeric cytokine (p70) which plays akey role in immune responses by bridging innate resistance andantigen-specific adaptive immunity. Trinehieri (1993) Immunol Today 14:335. For example, it promotes type 1 T helper cell (Th1) responses and,hence, cell-mediated immunity. (See Chan et al. (1991) J Exp Med 173:869; Seder et al. (1993) Proc Natl Acad Sci USA 90: 10188; Manetti etal. (1993) J Exp Med 177: 1199; and Hsieh et al. (1993) Science 260:547, the entire teachings of each of these references are incorporatedhere in by reference). IL-12 is composed of two, disulfide linked,independently regulated subunits, p35 and p40. IL-12 is produced byphagocytic cells and antigen presenting cells, in particular,macrophages and dendritic cells, upon stimulation with bacteria,bacterial products such as lipopolysaccharide (LPS), and intracellularparasites. The well-documented biological functions of IL-12 areinduction of interferon-γ (INF-γ) expression from T and NK cells anddifferentiation of naïve T cells toward the Th1 T lymphocyte type.IFN-γ, expression of which is induced by IL-12, is a strong andselective enhancer of IL-12 production from monocytes and macrophages.

The cytokine IL-23 is a heterodimer composed of a p19 subunit and thesame p40 subunit as IL-12. Exposure of activated CD4⁺ T cells to IL-23causes them to develop into a novel T cell subset (Th_(IL-17) cells)characterized by the production of IL-17 (Langrish, et al.,Immunological Reviews (2004), 202:96-105, the entire teachings of whichare incorporated herein by reference). IL-17 has been implicated in anumber of autoimmune diseases. For example, IL-23 knock out mice have aseverely impaired IL-17 response and are resistant to experimentalautoimmune encephalitis (EAE) (a model for multiple sclerosis) andcollagen-induced arthritis. In addition, IL-17 has been shown toincrease neutrophil activity in the lungs of patients with acute andchronic asthma and in patients with chronic obstructive pulmonarydisease (COPD).

IL-27 is formed by the association of EBI3, a polypeptide related to thep40 subunit of IL-12, and p28, a protein related to the p35 subunit ofIL-12. IL-27 promotes the growth of T cells and, like IL-12, is thoughtto play a role in the differentiation of naïve T cells to Th1 cells.Pflanz et al., Immunity (2002), 16:779-790.

It has been suggested that, particularly in chronic diseases in whichthere is ongoing production of IFN-γ, IL-12 production is augmented byIFN-γ. It is presumed that after an infective or inflammatory stimulusthat provokes IL-12 production, the powerful feedback loop promotesIL-12-induced IFN-γ to further augment IL-12 production, leading toconsequent excessive production of pro-inflammatory cytokines. Inparticular, activated CD4⁺ T cells produced by this process when exposedto IL-23 differentiate into a subset of T cells that produce thepro-inflammatory cytokine, IL-17. Furthermore, it has been suggestedthat IL-27 induces the expression of T-bet, a major Th1-specifictranscription factor, and its downstream target IL-12R β2, independentlyof IFN-γ. In addition, IL-27 suppresses the expression of GATA-3. GATA-3inhibits Th1 development and causes loss of IL-12 signaling throughsuppression of IL-12R β2 and Stat4 expression. Lucas et al., PNAS(2003), 100:15047-15052.

IL-12, IL-23 and IL-27, as well as IL-17, play a critical role in manyautoimmune diseases including, but not limited to, multiple sclerosis,sepsis, myasthenia gravis, autoimmune neuropathies, Guillain-Barrésyndrome, autoimmune uveitis, autoimmune hemolytic anemia, perniciousanemia, autoimmune thrombocytopenia, temporal arteritis,anti-phospholipid syndrome, vasculitides, Wegener's granulomatosis,Behcet's disease, psoriasis, psoriatic arthritis, dermatitisherpetiformis, pemphigus vulgaris, vitiligo, Crohn's disease, ulcerativecolitis, interstitial pulmonary fibrosis, myelofibrosis, hepaticfibrosis, myocarditis, thyroditis, primary biliary cirrhosis, autoimmunehepatitis, Type I or immune-mediated diabetes mellitus, Grave's disease,Hashimoto's thyroiditis, autoimmune oophoritis and orchitis, autoimmunedisease of the adrenal gland; rheumatoid arthritis, juvenile rheumatoidarthritis, systemic lupus erythematosus, scleroderma, polymyositis,dermatomyositis, spondyloarthropathies, ankylosing spondylitis,Sjogren's syndrome and graft-versus-host disease. (See, for example,Gately et al. (1998) Annu Rev Immunol. 16: 495; and Abbas et al. (1996)Nature 383: 787, the entire teachings of which are incorporated byreference.)

Inhibiting IL-12, IL-23 and IL-27 is an approach to treating autoimmuneand inflammatory disorders by inhibiting the production of Th1 andTh_(IL-17) cells and thereby down-regulating pro-inflammatory cytokinessuch as IL-17. (Trembleau et al. (1995) Immmunol. Today 16: 383; andAdorini et al. (1997) Chem. Immunol. 68: 175; and Langrish, et al.,Immunological Reviews (2004), 202:96-105, the entire teachings of bothof these articles are incorporated herein by reference). Therefore,compounds that inhibit the production of IL-12, IL-23 and IL-27 areuseful for treating autoimmune and inflammatory disorders.

SUMMARY

The present invention relates to compounds that inhibit the productionof IL-12, IL-23, and IL-27 and are useful in treating autoimmune andinflammatory disorders. In one aspect, this invention featurespyrimidine compounds of formula (I):

R₁ is

[referred to hereinafter as NC(R^(a)R^(b))], aryl, or heteroaryl; eachof R₂ and R₄, independently, is R^(c), halogen, nitro, cyano,isothionitro, SR^(c), or OR^(c); or R₂ and R₄, taken together, iscarbonyl; R₃ is R^(c), alkenyl, alkynyl, OR^(c), OC(O)R^(c), SO₂R^(c),S(O)R^(c), S(O₂)NR^(c)R^(d), SR^(c), NR^(c)R^(d), NR^(c)COR^(d),NR^(c)C(O)OR^(d), NR^(c)C(O)NR^(c)R^(d), NR^(c)SO₂R^(d), COR^(c),C(O)OR^(c), or C(O)NR^(c)R^(d); R₅ is H or alkyl; n is 0, 1, 2, 3, 4, 5,or 6; X is O, S, S(O), S(O₂), or NR^(c); Y is a covalent bond, CH₂,C(O), C═N—R^(c), C═N—OR^(c), C═N—SR^(c), O, S, S(O), S(O₂), or NR^(c); Zis N or CH; one of U and V is N, and the other is CR^(c); and W is O, S,S(O), S(O₂), NR^(c), or NC(O)R^(c); in which each of R^(a) and R^(b),independently, is H, alkyl, aryl, heteroaryl; and each of R^(c) andR^(d), independently, is H, alkyl, aryl, heteroaryl, cyclyl,heterocyclyl, or alkylcarbonyl. Also note that when n is 2 or greater,the just-described heteroaryl compound may have two or more differentC(R²R⁴) moieties. The same rule applies to other similar situations.

Referring to formula (I), a subset of the pyrimidine compounds of thisinvention is featured by that R¹ is NC(R^(a)R^(b)). In these compounds,U can be N, V can be CH, Z can be N, and W can be O. In addition, X canbe NR^(c); R^(c) can be H, methyl, ethyl, or acetyl; Y can be O or CH₂,and n can be 0, 1, 2, 3, or 4. In some embodiments, R₃ is aryl,heteroaryl (e.g., pyridinyl), OR^(c), SR^(c), C(O)OR^(c), orC(O)NR^(c)R^(d). In other embodiments, R₃ is

in which each of A and A′, independently, is O, S, or NH; each of R^(e)and R^(f), independently, is H, alkyl, aryl, or heteroaryl; and m is 1or 2.

In this subset of pyrimidine compounds, R^(a) or R^(b), preferably, is

in which B is NR^(i), O, or S; B′ is N or CR^(i); R^(g) is H, alkyl, oralkoxyl; R^(h) is halogen, NO₂, CN, alkyl, aryl, heteroaryl, OR^(c),OC(O)R^(c), SO₂R^(c), S(O)R^(c), S(O₂)NR^(c)R^(d), SR^(c), NR^(c)R^(d),NR^(c)COR^(d), NR^(c)C(O)OR^(d), NR^(c)C(O)NR^(c)R^(d), NR^(c)SO₂R^(d),COR^(c), C(O)OR^(c), or C(O)NR^(c)R^(d); R^(i) is H, alkyl, oralkylcarbonyl; p is 0, 1, or 2; and q is 0, 1, 2, 3, or 4. Preferably, Bis NR^(i); B′ is CH; R^(g) is H, methyl, ethyl, propyl, cyclopropyl,methoxy, or ethoxy; R^(h) is F, Cl, CN, methyl, methoxy, ethoxy,OC(O)CH₃, OC(O)C₂H₅, C(O)OH, C(O)OC₂H₅, C(O)NH₂, NHC(O)CH₃, or S(O₂)NH₂;R^(i) is H, methyl, ethyl, or acetyl; and q is 0, 1, or 2.

Another subset of the pyrimidine compounds of this invention is featuredby that R^(l) is aryl or heteroaryl. In these compounds, U can be N, Vcan be CH, Z can be N, and W can be O. In addition, X can be NR^(c);R^(c) can be H, methyl, ethyl, or acetyl; Y can be O or CH₂, and n canbe 0, 1, 2, 3, or 4. In some embodiments, R₃ is aryl, heteroaryl (e.g.,pyridinyl), OR^(c), SR^(c), C(O)OR^(c), or C(O)NR^(c)R^(d). In otherembodiments, R₃ is

in which each of A and A′, independently, is O, S, or NH; each of R^(e)and R^(f), independently, is H, alkyl, aryl or heteroaryl; and m is 1 or2.

In this second subset of pyrimidine compounds, R_(t), preferably, is

in which D is O, S, or NR^(m), R^(j) is benzo, halogen, CN, hydroxyl,alkyl, aryl, heteroaryl, alkoxyl, aryloxyl, or heteroaryloxyl; R^(m) isH, alkyl, or alkylcarbonyl; and r is 0, 1, or 2. Preferably, R₁ is

and R^(j) is methyl, ethyl, propyl, or benzo; and r can be 1 or 2.

Set forth below are exemplary compounds of this invention:

In another aspect, this invention features a pharmaceutical compositionthat contains a pharmaceutically acceptable carrier and an effectiveamount of at least one of the pyrimidine compounds of this invention, orpharmaceutically acceptable salt, solvate, clathrate, hydrate,polymorph, or prodrug thereof.

In another aspect, the present invention features a method for treatingor preventing an autoimmune or inflammatory disorder in a subject,wherein the autoimmune or inflammatory disorder is selected from thegroup consisting of ankylosing spondilitis, gastric ulcer, ulcerativecolitis, seronegative arthropathies, systemic lupus erythematosis,antiphospholipid syndrome, iridocyclitis/uveitis/optic neuritis,idiopathic pulmonary fibrosis, inflammatory pulmonary syndrome, systemicvasculitis/wegener's granulomatosis, sarcoidosis, orchitis/vasectomyreversal procedures, allergic/atopic diseases, asthma, allergicrhinitis, eczema, allergic contact dermatitis, allergic conjunctivitis,hypersensitivity pneumonitis, transplants, organ transplant rejection,graft-versus-host disease, systemic inflammatory response syndrome,neutropenic fever, urosepsis, meningococcemia, trauma/hemorrhage, burns,ionizing radiation exposure, acute pancreatitis, adult respiratorydistress syndrome, alcohol-induced hepatitis, chronic inflammatorypathologies, sarcoidosis, sickle cell anemia, type II diabetes,nephrosis, atopic diseases, hypersensitity reactions, allergic rhinitis,hay fever, perennial rhinitis, conjunctivitis, endometriosis, asthma,urticaria, systemic anaphalaxis, dermatitis, pernicious anemia,hemolytic disease, thrombocytopenia, graft rejection of any organ ortissue, kidney transplant rejection, heart transplant rejection, livertransplant rejection, pancreas transplant rejection, lung transplantrejection, bone marrow transplant (BMT) rejection, skin allograftrejection, cartilage transplant rejection, bone graft rejection, smallbowel transplant rejection, fetal thymus implant rejection, parathyroidtransplant rejection, xenograft rejection of any organ or tissue,allograft rejection, anti-receptor hypersensitivity reactions, Gravesdisease, Raynoud's disease, asthma, myasthenia gravis,antibody-meditated cytotoxicity, type III hypersensitivity reactions,POEMS syndrome (polyneuropathy, organomegaly, endocrinopathy, monoclonalgammopathy, and skin changes syndrome), polyneuropathy, organomegaly,endocrinopathy, monoclonal gammopathy, skin changes syndrome,antiphospholipid syndrome, pemphigus, scleroderma, mixed connectivetissue disease, idiopathic Addison's disease, chronic active hepatitis,primary billiary cirrhosis, vitiligo, vasculitis, post-MI cardiotomysyndrome, type IV hypersensitivity, contact dermatitis, hypersensitivitypneumonitis, allograft rejection, granulomas due to intracellularorganisms, drug sensitivity, metabolic/idiopathic, Wilson's disease,hemachromatosis, alpha-1-antitrypsin deficiency, diabetic retinopathy,hashimoto's thyroiditis, hypothalamic-pituitary-adrenal axis evaluation,primary biliary cirrhosis, thyroiditis, encephalomyelitis, cachexia,cystic fibrosis, neonatal chronic lung disease, chronic obstructivepulmonary disease (COPD), familial hematophagocytic lymphohistiocytosis,dermatologic conditions, alopecia, nephrotic syndrome, nephritis,glomerular nephritis, acute renal failure, hemodialysis, uremia,toxicity, preeclampsia, pemphigus vulgaris, chronic salicylateintoxication, idiopathic thrombocytopenic purpura, autoimmunemeningitis, myasthenia gravis, autoimmune thyroiditis, Sjogren'sSyndrome, alopecia areata, allergic responses due to arthropod bitereactions, aphthous ulcer, iritis, conjunctivitis, keratoconjunctivitis,cutaneous lupus erythematosus, scleroderma, vaginitis, proctitis, drugeruptions, leprosy reversal reactions, erythema nodosum leprosum,autoimmune uveitis, allergic encephalomyelitis, aplastic anemia, purered cell anemia, idiopathic thrombocytopenia, polychondritis, Wegener'sgranulomatosis, chronic active hepatitis, Graves ophthalmopathy, primarybiliary cirrhosis, uveitis posterior and interstitial lung fibrosis. Themethod comprises administering to a subject in need thereof a compoundof formula I, or any formula disclosed herein, or pharmaceuticallyacceptable salt, solvate, clathrate, hydrate, polymorph, or prodrugthereof.

In another aspect, the present invention features a method for treatingor preventing an autoimmune or inflammatory disorder in a subject,wherein the autoimmune or inflammatory disorder is selected from thegroup consisting of juvenile rheumatoid arthritis, systemic onsetjuvenile rheumatoid arthritis, psoriatic arthritis, ankylosingspondilitis, osteoarthritis, and Lyme arthritis. The method comprisesadministering to a subject in need thereof a compound of formula I, oran formula disclosed herein, or pharmaceutically acceptable salt,solvate, clathrate, hydrate, polymorph, or prodrug thereof.

In another aspect, the present invention features a method for treatingor preventing an autoimmune or inflammatory disorder in a subject,wherein the autoimmune or inflammatory disorder is selected from thegroup consisting of asthma, neonatal chronic lung disease, or chronicobstructive pulmonary disease. The method comprises administering to asubject in need thereof a compound of formula I, or any formuladisclosed herein, or pharmaceutically acceptable salt, solvate,clathrate, hydrate, polymorph, or prodrug thereof. As disclosed above,IL-23 stimulates activated CD4⁺ T cells to develop into a novel T cellsubset characterized by the production of IL-17. 1L-17 has been shown toincrease neutrophil activity in the lungs in patients with severe asthmaand in patients with chronic obstructive pulmonary disease.

In one embodiment, this invention features a method of inhibiting IL-23,IL-27, and/or IL-12 production in a subject, comprising administering tothe subject an effective amount of a compound of formula I, or anyformula disclosed herein, or pharmaceutically acceptable salt, solvate,clathrate, hydrate, polymorph, or prodrug thereof.

In another embodiment, the invention features a method of inhibitingdevelopment or proliferation of Th_(IL-17) cells, comprisingadministering to a subject in need thereof a compound of formula I, orany formula disclosed herein, or pharmaceutically acceptable salt,solvate, clathrate, hydrate, polymorph, or prodrug thereof. In oneembodiment, the method further comprises inhibiting the production ofIL-17.

In another embodiment, the invention features a method of inhibitingdevelopment or proliferation of Th1 cells, comprising administering to asubject in need thereof a compound of formula I, or any formuladisclosed herein, or pharmaceutically acceptable salt, solvate,clathrate, hydrate, polymorph, or prodrug thereof. In one embodiment,the method further comprises inhibiting the production of IL-17.

In addition, some of the pyrimidine compounds of this invention have oneor more double bonds, or one or more asymmetric centers. Such compoundscan occur as racemates, racemic mixtures, single enantiomers, individualdiastereomers, diastereomeric mixtures, and cis- or trans- or E- orZ-double isomeric forms. All such forms are included in the invention.

The details of one or more embodiments of the invention are set forth inthe accompanying drawings and the description below. Other features,objects, and advantages of the invention will be apparent from thedescription and drawings, and from the claims.

DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph showing the percent inhibition of IL-12, IL-23 and thep40 subunit of IL-12 and IL-23 by Compound 12 in human peripheral bloodmononuclear cells after stimulation with S. aureus Cowan I (SAC) in thepresence of IFN-γ.

FIG. 2 is a graph showing the percent inhibition of IL-12, IL-23 and thep40 subunit of IL-12 and IL-23 by Compound 12 in human peripheral bloodmononuclear cells after stimulation with liposaccharide (LPS) in thepresence of IFN-γ.

DETAILED DESCRIPTION

Alkyl, alkenyl, alkynyl, aryl, heteroaryl (e.g., pyridinyl), cyclyl,heterocyclyl mentioned above include both substituted and unsubstitutedmoieties. The term “substituted” refers to one or more substituents(which may be the same or different), each replacing a hydrogen atom.Examples of substituents include, but are not limited to, halogen,hydroxyl, amino, alkylamino, arylamino, dialkylamino, diarylamino,cyano, nitro, mercapto, alkylcarbonyl, carbamido, carbamyl, carboxyl,thioureido, thiocyanato, sulfoamido, C₁˜C₆ alkyl, C₁˜C₆ alkenyl, C₁˜C₆alkoxy, aryl, heteroaryl, cyclyl, heterocyclyl, wherein alkyl, alkenyl,alkoxy, aryl, heteroaryl, cyclyl, and heterocyclyl are optionallysubstituted with C₁˜C₆ alkyl, aryl, heteroaryl, halogen, hydroxyl,amino, mercapto, cyano, or nitro.

As used herein, the term “alkyl” refers to a straight-chained orbranched hydrocarbon group containing 1 to 12 carbon atoms. The term“lower alkyl” refers to a C1-C6 alkyl chain. Examples of alkyl groupsinclude methyl, ethyl, n-propyl, isopropyl, tert-butyl, and n-pentyl.Alkyl groups may be optionally substituted with one or moresubstituents.

The term “alkenyl” refers to an unsaturated hydrocarbon chain that maybe a straight chain or branched chain, containing 2 to 12 carbon atomsand at least one carbon-carbon double bond. Alkenyl groups may beoptionally substituted with one or more substituents.

The term “alkynyl” refers to an unsaturated hydrocarbon chain that maybe a straight chain or branched chain, containing the 2 to 12 carbonatoms and at least one carbon-carbon triple bond. Alkynyl groups may beoptionally substituted with one or more substituents.

The sp² or sp carbons of an alkenyl group and an alkynyl group,respectively, may optionally be the point of attachment of the alkenylor alkynyl groups.

The term “cyclyl” refers to a hydrocarbon 3-8 membered monocycle or 7-14membered bicycle ring system having at least one non-aromatic ring whichmay optionally have some degree of saturation. Cyclyl groups may beoptionally substituted with one or more substituents. In one embodiment,0, 1, 2, 3, or 4 atoms of each ring of a cyclyl group may be substitutedby a substituent. Representative examples of cyclyl group includecyclopropyl, cyclopentyl, cyclohexyl, cyclobutyl, cycloheptyl,cyclooctyl, cyclononyl, cyclodecyl, cyclohexenyl,bicyclo[2.2.1]hept-2-enyl, dihydronaphthalenyl, benzocyclopentyl,cyclopentenyl, cyclopentadienyl, cyclohexenyl, cyclohexadienyl,cycloheptenyl, cycloheptadienyl, cycloheptatrienyl, cyclooctenyl,cyclooctadienyl, cyclooctatrienyl, cyclooctatetraenyl, cyclononenyl,cyclononadienyl, cyclodecenyl, cyclodecadienyl and the like.

The term “aryl” refers to a hydrocarbon ring system having at least onearomatic ring. Typically, a monocycle aryl will have from 5 to 8 carbonatom ring members; a bicyclic ary will have from 7 to 14 carbon atomring members, and a tricyclic aryl will have 11-14 carbon atom ringmembers. Examples of aryl moieties include, but are not limited to,phenyl, naphthyl, anthracenyl, fluorenyl, indenyl, azulenyl, pyrenyl,and the like.

The term “heterocyclyl” refers to a nonaromatic 3-8 membered monocycle,7-12 membered bicyclic, or 10-14 membered tricyclic ring systemcomprising 1-3 heteroatoms if monocycle, 1-6 heteroatoms if bicyclic, or1-9 heteroatoms if tricyclic, said heteroatoms selected from O, N, S, B,P or Si, wherein the nonaromatic ring system may have some degree ofsaturation. Heterocyclyl groups may be optionally substituted with oneor more substituents. In one embodiment, 0, 1, 2, 3, or 4 atoms of eachring of a heterocyclyl group may be substituted by a substituent.Representative heterocyclyl groups include piperidinyl, piperazinyl,2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, 4-piperidonyl,tetrahydropyranyl, tetrahydrothiopyranyl, tetrahydrothiopyranyl sulfone,morpholinyl, thiomorpholinyl, thiomorpholinyl sulfoxide, thiomorpholinylsulfone, 1,3-dioxolane, tetrahydrofuranyl, tetrahydrothienyl, thiirenyl,thiadiazirinyl, dioxazolyl, 1,3-oxathiolyl, 1,3-dioxolyl, 1,3-dithiolyl,oxathiazinyl, dioxazinyl, dithiazinyl, oxadiazinyl, thiadiazinyl,oxazinyl, thiazinyl, 1,4-oxathiin,1,4-dioxin, 1,4-dithiin, 1H-pyranyl,oxathiepinyl, 5H-1,4-dioxepinyl, 5H-1,4-dithiepinyl,6H-isoxazolo[2,3-d]1,2,4-oxadiazolyl,7aH-oxazolo[3,2-d]1,2,4-oxadiazolyl, and the like.

The term “heteroaryl” refers to a hydrocarbon ring system having atleast one aromatic ring which contains at least one heteroatom such asO, N, or S. Typically, a monocyclic heteroaryl has 5-8 ring members, abicyclic heteroaryl has 7-12 ring members, and a tricyclic heteroarylhas 11-14 membered tricyclic ring system having 1-4 ring heteroatoms ifmonocyclic, 1-6 heteroatoms if bicyclic, or 1-9 heteroatoms iftricyclic, said heteroatoms selected from O, N, or S, and the remainderring atoms being carbon (with appropriate hydrogen atoms unlessotherwise indicated). Heteroaryl groups may be optionally substitutedwith one or more substituents. In one embodiment, 0, 1, 2, 3, or 4 atomsof each ring of a heteroaryl group may be substituted by a substituent.Examples of heteroaryl groups include pyridyl, 1-oxo-pyridyl, (uranyl,benzo[1,3]dioxolyl, benzo[1,4]dioxinyl, thienyl, pyrrolyl, oxazolyl,oxadiazolyl, imidazolyl thiazolyl, isoxazolyl, quinolinyl, pyrazolyl,isothiazolyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, triazolyl,thiadiazolyl, isoquinolinyl, indazolyl, benzoxazolyl, benzofuryl,indolizinyl, imidazopyridyl, tetrazolyl, benzimidazolyl, benzothiazolyl,benzothiadiazolyl, benzoxadiazolyl, indolyl, tetrahydroindolyl,azaindolyl, imidazopyridyl, quinazolinyl, purinyl,pyrrolo[2,3]pyrimidinyl, pyrazolo[3,4]pyrimidinyl, and benzo(b)thienyl,3H-thiazolo[2,3-c][1,2,4]thiadiazolyl,imidazo[1,2-b]-1,2,4-thiadiazolyl, imidazo[2,1-b]-1,3,4-thiadiazolyl,1H,2H-furo[3,4-d]-1,2,3-thiadiazolyl,1H-pyrazolo[5,1-c]-1,2,4-triazolyl, pyrrolo[3,4-d]-1,2,3-triazolyl,cyclopentatriazolyl, 3H-pyrrolo[3,4-c]isoxazolyl,1H,3H-pyrrolo[1,2-c]oxazolyl, pyrrolo[2,1b]oxazolyl, and the like.Examples of heteroaryl moieties include, but are not limited to, furyl,fluorenyl, thienyl, oxazolyl, imidazolyl, thiazolyl, pyridinyl,pyrimidinyl, quinazolinyl, and indolyl.

The compounds of this invention include the compounds themselves, aswell as their salts, solvate, clathrate, hydrate, polymorph, orprodrugs, if applicable. As used herein, the term “pharmaceuticallyacceptable salt,” is a salt formed from, for example, an acid and abasic group of a compound of any one of the formulae disclosed herein.Illustrative salts include, but are not limited, to sulfate, citrate,acetate, oxalate, chloride, bromide, iodide, nitrate, bisulfate,phosphate, acid phosphate, isonicotinate, lactate, salicylate, acidcitrate, tartrate, oleate, tannate, pantothenate, bitartrate, ascorbate,succinate, maleate, besylate, gentisinate, fumarate, gluconate,glucaronate, saccharate, formate, benzoate, glutamate, methanesulfonate,ethanesulfonate, benzenesulfonate, p-toluenesulfonate, and pamoate(i.e., 1,1′-methylene-bis-(2-hydroxy-3-naphthoate)) salts. The term“pharmaceutically acceptable salt” also refers to a salt prepared from acompound of any one of the formulae disclosed herein having an acidicfunctional group, such as a carboxylic acid functional group, and apharmaceutically acceptable inorganic or organic base. Suitable basesinclude, but are not limited to, hydroxides of alkali metals such assodium, potassium, and lithium; hydroxides of alkaline earth metal suchas calcium and magnesium; hydroxides of other metals, such as aluminumand zinc; ammonia, and organic amines, such as unsubstituted orhydroxy-substituted mono-, di-, or trialkylamines; dicyclohexylamine;tributyl amine; pyridine; N-methyl, N-ethylamine; diethylamine;diethylamine; mono-, bis-, or tris-(2-hydroxy-lower alkyl amines), suchas mono-, bis-, or tris-(2-hydroxyethyl)amine,2-hydroxy-tert-butylamine, or tris-(hydroxymethyl)methylamine,N,N,-di-lower alkyl-N-(hydroxy lower alkyl)-amines, such asN,N-dimethyl-N-(2-hydroxyethyl)amine, or tri-(2-hydroxyethyl)amine;N-methyl-D-glucamine; and amino acids such as arginine, lysine, and thelike. The term “pharmaceutically acceptable salt” also refers to a saltprepared from a compound of any one of the formulae disclosed hereinhaving a basic functional group, such as an amino functional group, anda pharmaceutically acceptable inorganic or organic acid, Suitable acidsinclude hydrogen sulfate, citric acid, acetic acid, oxalic acid,hydrochloric acid (HCl), hydrogen bromide (HBr), hydrogen iodide (HI),nitric acid, hydrogen bisulfide, phosphoric acid, lactic acid, salicylicacid, tartaric acid, bitartratic acid, ascorbic acid, succinic acid,maleic acid, besylic acid, fumaric acid, gluconic acid, glucaronic acid,formic acid, benzoic acid, glutamic acid, methanesulfonic acid,ethanesulfonic acid, benzenesulfonic acid, and p-toluenesulfonic acid.

As used herein, the term “polymorph” means solid crystalline forms of acompound of the present invention or complex thereof. Differentpolymorphs of the same compound can exhibit different physical, chemicaland/or spectroscopic properties. Different physical properties include,but are not limited to stability (e.g., to heat or light),compressibility and density (important in formulation and productmanufacturing), and dissolution rates (which can affectbioavailability). Differences in stability can result from changes inchemical reactivity (e.g., differential oxidation, such that a dosageform discolors more rapidly when comprised of one polymorph than whencomprised of another polymorph) or mechanical characteristics (e.g.,tablets crumble on storage as a kinetically favored polymorph convertsto thermodynamically more stable polymorph) or both (e.g., tablets ofone polymorph are more susceptible to breakdown at high humidity).Different physical properties of polymorphs can affect their processing.For example, one polymorph might be more likely to form solvates ormight be more difficult to filter or wash free of impurities thananother due to, for example, the shape or size distribution of particlesof it.

As used herein, the term “hydrate” means a compound of the presentinvention or a salt thereof, which further includes a stoichiometric ornon-stoichiometric amount of water bound by non-covalent intermolecularforces.

As used herein, the term “clathrate” means a compound of the presentinvention or a salt thereof in the form of a crystal lattice thatcontains spaces (e.g., channels) that have a guest molecule (e.g., asolvent or water) trapped within.

As used herein and unless otherwise indicated, the term “prodrug” meansa derivative of a compound that can hydrolyze, oxidize, or otherwisereact under biological conditions (in vitro or in vivo) to provide acompound of this invention. Prodrugs may only become active upon suchreaction under biological conditions, or they may have activity in theirunreacted forms. Examples of prodrugs contemplated in this inventioninclude, but are not limited to, analogs or derivatives of compounds ofany one of the formulae disclosed herein that comprise biohydrolyzablemoieties such as biohydrolyzable amides, biohydrolyzable esters,biohydrolyzable carbamates, biohydrolyzable carbonates, biohydrolyzableureides, and biohydrolyzable phosphate analogues. Other examples ofprodrugs include derivatives of compounds of any one of the formulaedisclosed herein that comprise —NO, —NO₂, —ONO, or —ONO₂ moieties.Prodrugs can typically be prepared using well-known methods, such asthose described by 1 BURGER'S MEDICINAL CHEMISTRY AND DRUG DISCOVERY(1995) 172-178, 949-982 (Manfred E. Wolff ed., 5^(th) ed).

As used herein and unless otherwise indicated, the terms“biohydrolyzable amide”, “biohydrolyzable ester”, “biohydrolyzablecarbamate”, “biohydrolyzable carbonate”, “biohydrolyzable ureide” and“biohydrolyzable phosphate analogue” mean an amide, ester, carbamate,carbonate, ureide, or phosphate analogue, respectively, that either: 1)does not destroy the biological activity of the compound and confersupon that compound advantageous properties in vivo, such as uptake,duration of action, or onset of action; or 2) is itself biologicallyinactive but is converted in vivo to a biologically active compound.Examples of biohydrolyzable amides include, but are not limited to,lower alkyl amides, α-amino acid amides, alkoxyacyl amides, andalkylaminoalkylcarbonyl amides. Examples of biohydrolyzable estersinclude, but are not limited to, lower alkyl esters, alkoxyacyloxyesters, alkyl acylamino alkyl esters, and choline esters. Examples ofbiohydrolyzable carbamates include, but are not limited to, loweralkylamines, substituted ethylenediamines, aminoacids,hydroxyalkylamines, heterocyclic and heteroaromatic amines, andpolyether amines.

As used herein, the terms “animal”, “subject” and “patient”, include,but are not limited to, a cow, monkey, horse, sheep, pig, chicken,turkey, quail, cat, dog, mouse, rat, rabbit, guinea pig and human(preferably, a human).

As used herein, the term “a subject in need thereof” refers to a subjectsuffering from an autoimmune or inflammatory disorder or who has apredisposition (e.g., a genetic predisposition) to develop an autoimmuneor inflammatory disorder. In addition, subjects that have had anautoimmune or inflammatory disorder that is in remission may be in needof treatment with one or more compounds of the invention, or apharmaceutically acceptable salt, solvate, clathrate, hydrate,polymorph, or prodrug thereof, to prevent a relapse of the autoimmune orinflammatory disorder.

The compounds described above can be prepared by methods well known inthe art, as well as by the synthetic routes disclosed herein anddescribed in U.S. Pat. Nos. 6,693,097, 6,660,733, 6,858,606 and in U.S.Provisional Application No. 60/626,609, the entire teaching of each ofthese patents and patent application are incorporated herein byreference. For example, a pyrimidine compound (e,g., Compounds 1-27) canbe prepared by using 2,4,6-trichloro-pyrimidine as a starting material.The three chloro groups can be displaced by various substitutes. Morespecifically, first chloro group (e.g., at position 6) can react with,e.g., morpholine, to form a morpholinyl pyrimidine. 2-Aryl and2-alkylpyrimidinde dichloro compounds can also be prepared by reactingan amidine with a malonic ester followed by treatment with phosphorousoxychloride. Second chloro group can be replaced by reacting with anucleophile, such as an alcohol in the presence of base, e.g., sodiumhydride. In other examples, a compound of formula (I), wherein Y is CH₂(e.g., Compound 1, 3-5, 11, 14, and 27), can be prepared by reacting thepyrimidine chloride with a Grignard reagent, an organotin reagent, anorganocopper reagent, an organoboric acid, or an organozinc reagent inthe presence of an organopalladium compound as a catalyst. Isomericforms may be produced. The desired isomeric product can be separatedfrom others by, e.g., high performance liquid chromatography. Thirdchloro group undergoes a displacement reaction with, e.g., hydrazine,and the primary amine of the coupled hydrazine moiety further reactswith an aldehyde, e.g., indole-3-carboxaldehyde to form a hydrazonelinkage. Thus, a pyrimidine compound of this invention is obtained. Ifpreferred, other types of linkages can be prepared by similar reactions.Sensitive moieties on a pyrimidinyl intermediate and a nucleophile canbe protected prior to coupling. For suitable protecting groups, see,e.g., Greene (1981) Protective Groups in Organic Synthesis, John Wiley &Sons, Inc., New York, the entire teachings of which are incorporatedherein by reference. A pyrimidine compound of this invention can befurther purified by flash column chromatography, high performance liquidchromatography, or crystallization.

The compounds and compositions described herein are useful to treat andprevent any inflammatory and immune disorders. In particular, thecompounds of the invention are useful in inhibiting the production ofIL-12, IL-23 and/or IL-27. IL-12 and IL-27 produce INF-γ which furtheraugments the production of IL-12 and causes the differentiation of naïveT cells into T_(H)1 lymphocytes which have been implicated in thepathogenic processes of many autoimmune and inflammatory disorders.IL-23 has been shown to stimulate the differentiation of activated CD4⁺T cells into Th_(IL-17) cells which produce IL-17, another cytokine thathas been implicated in the pathogenic processes of many autoimmune andinflammatory disorders. Thus, in one aspect, the present inventionprovides a method of treating or preventing autoimmune or inflammatorydisorders by inhibiting the production of IL-12, IL-27 and/or IL-23 in asubject by administering to the subject in need thereof an effectiveamount of a compound of any of the formulae herein, or apharmaceutically acceptable salt, solvate, clathrate, hydrate,polymorph, or prodrug thereof.

Without wishing to be bound by any theory, since one of the functions ofIL-12 and IL-27 is induction of INF-γ expression from T and NK cellswhich promotes the development of Th1 T lymphocyte type, the compoundsof the invention can be used to inhibit the differentiation of naïve Tcells into Th1 lymphocytes and/or inhibit the proliferation of Th1cells. Therefore, in another aspect, the invention features a method ofinhibiting the proliferation and/or development of Th1 cells in asubject in need thereof by administering to the subject an effectiveamount of a compound of any of the formulae herein, or apharmaceutically acceptable salt, solvate, clathrate, hydrate,polymorph, or prodrug thereof.

Without wishing to be bound by any theory, since one of the functions ofIL-23 is to promote the differentiation of activated CD4⁺ T cells toTh_(IL-17) lymphocyte type which produces the pro-inflammatory cytokineIL-17, the compounds of the invention can be used to inhibit thedifferentiation of activated CD4⁺ T cells into Th_(IL-17) lymphocytesand/or inhibit the proliferation of Th_(IL-17) lymphocytes. Therefore,in another aspect, the invention features a method of inhibiting theproliferation and/or development of Th_(IL-17) lymphocytes in a subjectin need thereof by administering to the subject an effective amount of acompound of any of the formulae herein, or a pharmaceutically acceptablesalt, solvate, clathrate, hydrate, polymorph, or prodrug thereof.

The term “inflammatory disorders” includes any inflammatory disease,disorder or condition caused, exasperated or mediated by IL-12, IL-23,IL-27 and/or IL-17 production. Such inflammatory disorders may include,without limitation, asthma, adult respiratory distress syndrome,systemic lupus erythematosus, inflammatory bowel disease (includingCrohn's disease and ulcerative colitis), multiple sclerosis,insulin-dependent diabetes mellitus, autoimmune arthritis (includingrheumatoid arthritis, juvenile rheumatoid arthritis, psoriaticarthritis), inflammatory pulmonary syndrome, pemphigus vulgaris,idiopathic thrombocytopenic purpura, autoimmune meningitis, myastheniagravis, autoimmune thyroiditis, dermatitis (including atopic dermatitisand eczematous dermatitis), psoriasis, Sjogren's Syndrome (includingkeratoconjunctivitis sicca secondary to Sjogren's Syndrome), alopeciaareata, allergic responses due to arthropod bite reactions, aphthousulcer, iritis, conjunctivitis, keratoconjunctivitis, cutaneous lupuserythematosus, scleroderma, vaginitis, proctitis, drug eruptions (suchas Stevens-Johnson syndrome), leprosy reversal reactions, erythemanodosum leprosum, autoimmune uveitis, allergic encephalomyelitis,aplastic anemia, pure red cell anemia, idiopathic thrombocytopenia,polychondritis, Wegener's granulomatosis, chronic active hepatitis,Graves ophthalmopathy, primary biliary cirrhosis, uveitis posterior andinterstitial lung fibrosis.

“Inflammatory disorders” expressly include acute inflammatory disorders.Examples of acute inflammatory disorders include graft versus hostdisease, transplant rejection, septic shock, endotoxemia, Lymearthritis, infectious meningitis (e.g., viral, bacterial, Lymedisease-associated), an acute episode of asthma and acute episodes of animmune disease.

“Inflammatory disorders” expressly include chronic inflammatorydisorders. Nonlimiting examples of chronic inflammatory disorder includeasthma, rubella arthritis, and chronic autoimmune diseases, such assystemic lupus erythematosus, psoriasis, inflammatory bowel disease,including Crohn's disease and ulcerative colitis, multiple sclerosis andrheumatoid arthritis.

The term “immune disorders” or “autoimmune disorders” includes anyimmune disease, disorder or condition caused, exasperated or mediated byIL-12, IL-23 and/or IL-27 production. Such immune diseases may include,without limitation, rheumatoid arthritis, juvenile rheumatoid arthritis,systemic onset juvenile rheumatoid arthritis, psoriatic arthritis,ankylosing spondilitis, gastric ulcer, seronegative arthropathies,osteoarthritis, inflammatory bowel disease, ulcerative colitis, systemiclupus erythematosis, antiphospholipid syndrome,iridocyclitis/uveitis/optic neuritis, idiopathic pulmonary fibrosis,systemic vasculitis/wegener's granulomatosis, sarcoidosis,orchitis/vasectomy reversal procedures, allergic/atopic diseases,asthma, allergic rhinitis, eczema, allergic contact dermatitis, allergicconjunctivitis, hypersensitivity pneumonitis, transplants, organtransplant rejection, graft-versus-host disease, systemic inflammatoryresponse syndrome, sepsis syndrome, gram positive sepsis, gram negativesepsis, culture negative sepsis, fungal sepsis, neutropenic fever,urosepsis, meningococcemia, trauma/hemorrhage, burns, ionizing radiationexposure, acute pancreatitis, adult respiratory distress syndrome,rheumatoid arthritis, alcohol-induced hepatitis, chronic inflammatorypathologies, sarcoidosis, Crohn's pathology, sickle cell anemia,diabetes, nephrosis, atopic diseases, hypersensitity reactions, allergicrhinitis, hay fever, perennial rhinitis, conjunctivitis, endometriosis,asthma, urticaria, systemic anaphalaxis, dermatitis, pernicious anemia,hemolytic disease, thrombocytopenia, graft rejection of any organ ortissue, kidney transplant rejection, heart transplant rejection, livertransplant rejection, pancreas transplant rejection, lung transplantrejection, bone marrow transplant (BMT) rejection, skin allograftrejection, cartilage transplant rejection, bone graft rejection, smallbowel transplant rejection, fetal thymus implant rejection, parathyroidtransplant rejection, xenograft rejection of any organ or tissue,allograft rejection, anti-receptor hypersensitivity reactions, Gravesdisease, Raynoud's disease, type B insulin-resistant diabetes, asthma,myasthenia gravis, antibody-meditated cytotoxicity, type IIIhypersensitivity reactions, systemic lupus erythematosus, POEMS syndrome(polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy,and skin changes syndrome), polyneuropathy, organomegaly,endocrinopathy, monoclonal gammopathy, skin changes syndrome,antiphospholipid syndrome, pemphigus, scleroderma, mixed connectivetissue disease, idiopathic Addison's disease, diabetes mellitus, chronicactive hepatitis, primary billiary cirrhosis, vitiligo, vasculitis,post-MI cardiotomy syndrome, type IV hypersensitivity, contactdermatitis, hypersensitivity pneumonitis, allograft rejection,granulomas due to intracellular organisms, drug sensitivity,metabolic/idiopathic, Wilson's disease, hemachromatosis,alpha-1-antitrypsin deficiency, diabetic retinopathy, hashimoto'sthyroiditis, osteoporosis, hypothalamic-pituitary-adrenal axisevaluation, primary biliary cirrhosis, thyroiditis, encephalomyelitis,cachexia, cystic fibrosis, neonatal chronic lung disease, chronicobstructive pulmonary disease (COPD), familial hematophagocyticlymphohistiocytosis, dermatologic conditions, psoriasis, alopecia,nephrotic syndrome, nephritis, glomerular nephritis, acute renalfailure, hemodialysis, uremia, toxicity, preeclampsia, okt3 therapy,anti-cd3 therapy, cytokine therapy, chemotherapy, radiation therapy(e.g., including but not limited to asthenia, anemia, cachexia, and thelike), chronic salicylate intoxication, and the like, See, e.g., theMerck Manual, 12th-17th Editions, Merck & Company, Rahway, N.J. (1972,1977, 1982, 1987, 1992, 1999), Pharmacotherapy Handbook, Wells et al.,eds., Second Edition, Appleton and Lange, Stamford, Conn. (1998, 2000),each entirely incorporated by reference.

The compounds and compositions described herein are useful to treat andprevent inflammatory disorders and immune disorders. The method involvesadministering an effective amount of a compound of the invention, or apharmaceutically acceptable salt, solvate, clathrate, hydrate,polymorph, or prodrug thereof, to a subject in need of treatment for aninflammatory or autoimmune disorder. In preferred embodiments, treatmentaccording to the invention provides a reduction in or prevention of atleast one symptom or manifestation of an IL-12-, IL-23-, IL-27, orIL-17-related disorder (e.g., inflammatory disorder or immune diseases),as determined in vivo or in vitro of at least about 10%, more preferably20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 98% or 99%.

As used herein, the term “effective amount” refers to an amount of acompound of this invention which is sufficient to reduce or amelioratethe severity, duration, progression, or onset of an inflammatorydisorder or immune disorder, or one or more symptom thereof, prevent theadvancement of an inflammatory disorder or immune disorder, cause theregression of an inflammatory disorder or immune disorder, prevent therecurrence, development, onset or progression of a symptom associatedwith an inflammatory disorder or immune disorder, or enhance or improvethe prophylactic or therapeutic effect(s) of another therapy. Aneffective amount of the pyrimidine compound of this invention can rangefrom about 0.001 mg/Kg to about 1000 mg/Kg. Effective doses will alsovary, as recognized by those skilled in the art, depending on thedisorder treated, route of administration, excipient usage, the age andsex of the subject, and the possibility of co-usage with othertherapeutic treatments such as use of other agents.

Also within the scope of this invention is a pharmaceutical compositionthat contains one or more pyrimidine compounds of this invention and apharmaceutically acceptable carrier.

To practice the method of the present invention, a pyrimidine compound,as a component of a pharmaceutical composition, can be administeredorally, parenterally, by inhalation spray, topically, rectally, nasally,buccally, vaginally or via an implanted reservoir. The term “parenteral”as used herein includes subcutaneous, intracutaneous, intravenous,intramuscular, intraarticular, intraarterial, intrasynovial,intrasternal, intrathecal, intralesional and intracranial injection orinfusion techniques.

A sterile injectable composition, for example, a sterile injectableaqueous or oleaginous suspension, can be formulated according totechniques known in the art using suitable dispersing or wetting agents(such as, for example, Tween 80) and suspending agents. The sterileinjectable preparation can also be a sterile injectable solution orsuspension in a non-toxic parenterally acceptable diluent or solvent,for example, as a solution in 1,3-butanediol. Among the acceptablevehicles and solvents that can be employed are mannitol, water, Ringer'ssolution and isotonic sodium chloride solution. In addition, sterile,fixed oils are conventionally employed as a solvent or suspending medium(e.g., synthetic mono- or diglycerides). Fatty acids, such as oleic acidand its glyceride derivatives are useful in the preparation ofinjectables, as are natural pharmaceutically-acceptable oils, such asolive oil or castor oil, especially in their polyoxyethylated versions.These oil solutions or suspensions can also contain a long-chain alcoholdiluent or dispersant, or carboxymethyl cellulose or similar dispersingagents. Other commonly used surfactants such as Tweens or Spans or othersimilar emulsifying agents or bioavailability enhancers which arecommonly used in the manufacture of pharmaceutically acceptable solid,liquid, or other dosage forms can also be used for the purposes offormulation.

A composition for oral administration can be any orally acceptabledosage form including, but not limited to, capsules, tablets, emulsionsand aqueous suspensions, dispersions and solutions. In the case oftablets for oral use, carriers which are commonly used include lactoseand corn starch. Lubricating agents, such as magnesium stearate, arealso typically added. For oral administration in a capsule form, usefuldiluents include lactose and dried corn starch. When aqueous suspensionsor emulsions are administered orally, the active ingredient can besuspended or dissolved in an oily phase combined with emulsifying orsuspending agents. If desired, certain sweetening, flavoring, orcoloring agents can be added. A nasal aerosol or inhalation compositioncan be prepared according to techniques well-known in the art ofpharmaceutical formulation and can be prepared as solutions in saline,employing benzyl alcohol or other suitable preservatives, absorptionpromoters to enhance bioavailability, fluorocarbons, and/or othersolubilizing or dispersing agents known in the art. A pyrimidinecompound of this invention can also be administered in the form ofsuppositories for rectal administration.

The carrier in the pharmaceutical composition must be “acceptable” inthe sense of being compatible with the active ingredient of theformulation (and preferably, capable of stabilizing it) and notdeleterious to the subject to be treated. For example, solubilizingagents such as cyclodextrins, which form specific, more solublecomplexes with the compounds of this invention, or one or moresolubilizing agents, can be utilized as pharmaceutical excipients fordelivery of the pyrimidine compounds. Examples of other carriers includecolloidal silicon dioxide, magnesium stearate, cellulose, sodium laurylsulfate, and D&C Yellow #10.

The biological activities of a pyrimidine compound can be evaluated by anumber of cell-based assays. One of such assays can be conducted usingcells from human peripheral blood mononuclear cells (PBMC) or humanmonocytic cell line (THP-1). The cells are stimulated with a combinationof human interferon-γ (IFNγ) and lipopolysaccharide or a combination ofIFNγ and Staphylococcus aureus Cowan I (SAC) in the presence of a testcompound. The level of inhibition of IL-12 production can be measured bydetermining the amount of p70 by using a sandwich ELISA assay withanti-human IL-12 antibodies. IC₅₀ of the test compound can then bedetermined. Specifically, PBMC or THP-1 cells are incubated with thetest compound. Cell viability is assessed using the bioreduction of MTS{3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium](Promega, Madison, Wis.).

The level of inhibition of IL-23 inhibition by a compound of theinvention can be measured by a similar assay in which human peripheralblood mononuclear cells (PBMC) or human monocytic cell line (THP-1) arestimulated with a combination of human interferon-γ (IFNγ) andlipopolysaccharide (LPS) or a combination of IFNγ and Staphylococcusaureus Cowan I (SAC) in the presence of a test compound. The level ofinhibition of IL-23 production can be measured by determining the amountof p19 by using a sandwich ELISA assay with antibodies the recognize p19subunit of IL-23. IC₅₀ of the test compound can then be determined. Onesuch assay is disclosed herein in Example 30.

The following specific embodiments are to be construed as merelyillustrative, and not limitative of the remainder of the disclosure inany way whatsoever. All of the patents, patent applications, andpublications cited herein are hereby incorporated by reference in theirentirety.

EXAMPLE 1 Preparation of Compound 1:N-{2-[3-(3,4-dimethoxy-phenyl)-propyl]-6-morpholin-4-yl-pyrimidin-4-yl}-N′-(1H-indol-3-ylmethylene)-hydrazine

To a solution of 3-(3,4-dimethoxyphenyl)-propyl iodide (1.224 g, 4.0mmol) in 20 mL dry THF, highly active zinc (suspension in THF, Riekemetal from Aldrich, 5.2 mL 0.05 g/mL, 4.0 mmol) was added to obtain amixture. The mixture was stirred at room temperature overnight.2,4-dichloro-6-morpholinopyrimidine (0.932 g, 4.0 mmol) andtrans-benzyl-(chloro)-bis-(triphenylphosphine)palladium(II) (0.03 g,0.04 mmol) were added to the mixture, and stirred at 60° C. for 2 days.After routine workup,4-chloro-2-[3-(3,4-dimethoxyphenyl)propyl]-6-morpholinopyrimidine (0.34g, 0.90 mmol, 22.4%) was separated from2-chloro-4-[3-(3,4-dimethoxyphenyl)propyl]-6-morpholinopyrimidine (0.45g, 1.19 mmol, 30%) by flash chromatography purification.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 6.70-6.80 (m, 3H); 6.32 (s, 1H); 3.87(s, 3H); 3.85 (s, 3H); 3.73-3.78 (m, 4H); 3.60-3.64 (m, 4H); 2.76 (d,J=7.8 Hz, 2H); 2.63 (d, J=7.5 Hz, 2H); and 2.01-2.12 (m, 2H).

MS (ESI): m/z 380.2 (M+H).

Further,4-chloro-2-[3-(3,4-dimethoxyphenyl)propyl]-6-morpholinopyrimidine (0.34g, 0.90 mmol) was reacted with hydrazine (0.29 g, 9 mmol) to obtain2-[3-(3,4-dimethoxyphenyl)propyl]-4-hydrazino-6-morpholinopyrimidine asa white solid (0.30 g, 0.80 mmol, 89%).

¹H NMR (300 MHz, CDCl₃), δ (ppm): 6.73-6.80 (m, 3H); 5.88 (s, 1H); 5.74(s, 1H); 3.87 (s, 3H); 3.85 (s, 3H); 3.76-3.79 (m, 4H); 3.69 (d, J=0.6Hz, 2H); 3.56-3.60 (m, 4H); 2.64 (d, J=7.5 Hz, 4H); and 2.00-2.15 (m,2H).

MS (ESI): m/z 374.2 (M−H).

A 5 mL methanol solution containing2-[3-(3,4-dimethoxyphenyl)-propyl]-4-hydrazino-6-morpholinopyrimidine(0.177 g, 0.50 mmol), indole-3-carboxaldehyde (0.073 g, 0.50 mmol), andAcOH (20 mg, cat.) was stirred at 70° C. for 4 hours. Solvent wasremoved and the crude residue was purified using flash chromatography togive Compound 1 as a light brown solid (0.21 g, 0.42 mmol, 84%).

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.57 (br s, 1H); 8.45 (br s, 1H);8.29-8.32 (m, 1H); 8.00 (s, 1H); 7.39-7.43 (m, 2H); 7.23-7.34 (m, 2H);6.74-6.80 (m, 3H); 6.30 (s, 1H); 3.86 (s, 3H); 3.85 (s, 3H); 3.78-3.84(m, 4H); 3.67-3.70 (m, 4H); 2.63-2.71 (m, 4H), and 2.03-2.13 (m, 2H).

MS (EST): m/z 501.2 (M+H).

EXAMPLE 2 Preparation of Compound 2:N-(2-n-butoxy-6-morpholin-4-yl-pyrimidin-4-yl)-N′-(1H-indol-3-ylmethylene)-hydrazine

To a solution of 2,4,6-trichloro pyrimidine (25 g, 136 mmol) in CH₂Cl₂(500 mL) at −78° C., morpholine (11.89 mL, 136 mmol) was slowly added,followed by DIPEA (25 mL, 143 mmol). The obtained reaction mixture wasstirred at −78° C. for 5 h, and then warmed up to room temperature. Thereaction mixture was washed with water. The obtained organic phase wasdried over Na₂SO₄. The solvent was removed under reduced pressure. Thecrude residue, 2,4-Dichloro-6-(morpholin-4-yl)pyrimidine, wasrecrystallized from EtOAc to give white crystals (24.7 g, 77%) 15 g.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 6.40 (s, 1H); and 4.0-3.5 (m, 8H).

MS (ESI): m/z 234.0 (M+H).

To a solution of n-butanol (0.633 g, 8.54 mmol) in anhydrous DMF (50 mL)at 0° C. under the N₂, NaH (0.307 g, 12.8 mmol) was added quickly. Theobtained suspension was stirred for 0.5 h at 0° C.2,4-Dichloro-6-(morpholin-4-yl)pyrimidine (2 g, 8.54 mmol) was added tothe suspension. After the suspension was warmed to room temperature andstirred for 12 h, the reaction mixture was quenched with ice/brine andextracted with 200 mL EtOAc. The extract was washed with brine, anddried over Na₂SO₄. The solvent was removed under reduced pressure. Thecrude residue was purified using flash chromatography (silica;EtOAc/Hexane: 1/6) to yield 1.4 g of2-n-butoxy-4-chloro-6-(morpholin-4-yl)pyrimidine (white solid, 60%).

¹H NMR (300 MHz, CDCl₃), δ (ppm): 6.20 (s, 1H); 4.26 (t, J=6.6 Hz, 2H);3.78-3.70 (m, 4H); 3.66-3.56 (m, 4H); 1.80-1.68 (m, 2H); 1.54-1.40 (m,2H); and 0.96 (t, 6.9, 3H).

MS (ESI): m/z 272.1 (M+H).

To a solution of 2-n-butoxy-4-chloro-6-(morpholin-4-yl)pyrimidine (1.38g, 5.1 mmol) in dioxane (50 ml), anhydrous hydrazine (1.6 mL, 50 mmol)was added. The obtained reaction mixture was heated to 95° C., andstirred for 12 h under N₂. After cooling to room temperature, thereaction mixture was quenched with ice-brine and extracted with EtOAc(200 mL). The organic extract was washed with brine, water, and driedover Na₂SO₄. The solvent was removed under reduced pressure. The cruderesidue was recrystallized from methanol to obtain2-n-butoxy-4-hydrazino-6-(morpholin-4-yl)pyrimidine as white crystals(1.10 g, 81%).

¹H NMR (300 MHz, CDCl₃), δ (ppm): 5.89 (br s, 1H), 5.49 (s, 1H), 4.26(t, J=6.6, 2H), 3.84-3.78 (m, 6H), 3.62-3.47 (m, 4H), 1.82-1.67 (m, 2H),1.55-1.42 (m, 2H), and 0.96 (t, J=6.9, 3H);

MS (ESI): m/z 268.2 (M+H).

To a solution of 2-n-butoxy-4-hydrazino-6-(morpholin-4-yl)pyrimidine(200 mg, 0.748 mmol) in MeOH (20 mL), indole-3-carboxaldehyde (108.6 mg,0.748 mmol) and acetic acid (a drop) were added sequentially. Theobtained reaction mixture was stirred at room temperature for 12 h.White precipitate was formed, collected, and washed with 2 mL methanolto give 200 g of Compound 2 (68%).

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.36 (br s, 1H), 8.30 (dd, J=6.6, 1.8,1H), 8.05 (s, 1H), 8.00 (s, 1H), 7.44-7.40 (m, 2H), 7.33-7.24 (m, 2H),6.13 (s, 1H), 4.26 (t, 2H, J=6.6), 3.84-3.78 (m, 4H), 3.70-3.64 (m, 4H),1.80-1.70 (m, 2H), 1.54-1.42 (m, 2H), and 0.96 (t, J=6.9, 3H);

MS (ESI): m/z 395.2 (M+H).

EXAMPLE 3 Preparation of Compound 3:N-(2-(4-hydroxybutyl)-6-morpholin-4-yl-pyrimidin-4-yl)-N′-(1H-indol-3-ylmethylene)-hydrazine

A mixture of 4-ethoxy-4-oxo-butylzinc bromide (50 mL 0.5M in THF, 25mmol), 2,4-dichloro-6-morpholinopyrimidine (4.68 g, 20.0 mmol) andtrans-benzyl(chloro)bis(triphenylphosphine)palladium(II) (0.15 g, 0.2mmol) in THF (total volume 80 mL) was stirred at 60° C. for 2 days.After routine workup, flash chromatography purification was performed toobtain 4-chloro-2-(4-ethoxy-4-oxo-butyl)-6-morpholinopyrimidine as awhite solid (2.073 g, 6.60 mmol, 33.0%).

To a solution of4-chloro-2-(4-ethoxy-4-oxo-butyl)-6-morpholinopyrimidine (1.108 g, 3.54mmol) in 50 mL THF at −78° C., a diisobutylaluminum hydride (DIBAL)solution (4.72 mL 1.5 M in Toluene, 7.08 mmol) was slowly added. Afteraddition, the obtained reaction mixture was warmed up slowly to 0° C.and kept at 0° C. for 10 min. After routine workup, flash chromatographywas performed to obtain4-chloro-2-(4-hydroxybutyl)-6-morpholinopyrimidine (0.76 g, 2.80 mmol,79%) as light yellow solid.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 6.33 (s, 1H), 3.76-3.79 (m, 4H);3.61-3.68 (m, 6H); 2.76 (t, J=7.8 Hz, 2H); 1.81-1.91 (m, 2H); and1.60-1.74 (m, 3H).

MS (ESI): m/z 370.2 (M+H).

Following the typical procedure,4-chloro-2-(4-hydroxybutyl)-6-morpholinopyrimidine (0.542 g, 2.00 mmol,1.00 equiv.) was reacted with hydrazine and indole-3-carboxaldehyde togive Compound 3 as an off-white solid (0.75 g, 1.90 mmol, 95%).

¹H NMR (300 MHz, DMSO-d₆), δ (ppm): 11.47 (s, 1H); 10.64 (s, 1H); 8.25(s, 1H); 8.18 (d, J=6.6 Hz, 1H); 7.71 (s, 1H); 7.43 (d, J=8.4 Hz, 1H);7.17-7.20 (m, 2H); 6.16 (s, 1H), 4.37 (t, J=4.8 Hz, 1H); 3.72 (br s,4H); 3.55 (br s, 4H); 3.41-3.45 (m, 2H); 2.49-2.54 (m, 2H), 1.66-1.76 (m2H); and 1.42-1.53 (m 2H).

MS (EST): m/z 395.1 (M+H).

EXAMPLE 4 Preparation of Compound 4:N-[2-(2-[1,3]dioxan-2-yl-ethyl)-6-morpholin-4-yl-pyrimidin-4-yl]-N′-(1H-indol-3-ylmethylene)-hydrazine

Compound 4 was prepared in a similar manner as described in Example 1.

¹H NMR (300 MHz, DMSO-d₆), δ (ppm): 11.46 (s, 1H); 10.64 (s, 1H); 8,25(s, 1H); 8.18 (d, J=6.6 Hz, 1H); 7.71 (s, 1H); 7.43 (d, J=6.0 Hz, 7.5Hz, 1H); 7.16-7.19 (m, 2H); 6.15 (s, 1H), 4.58 (t, J=5.1 Hz, 1H); 4.00(dd, J=11.4 Hz, 4.5 Hz, 2H); 3.64-3.72 (m, 6H); 3.54 (br s, 4H);2.50-2,59 (m, 2H); 1.80-1.94 (m, 3H), and 1.33 (d, J=9.6 Hz, 1H).

MS (EST): m/z 437.2 (M+H).

EXAMPLE 5 Preparation of Compound 5:N-(1H-indol-3-ylmethylene)-N′-[2-(3-methoxypropyl)-6-morpholin-4-yl-pyrimidin-4-yl]-hydrazine

Following the procedure for the synthesis ofN-(2-(4-Hydroxybutyl)-6-morpholin-4-yl-pyrimidin-4-yl)-N′-(1H-indol-3-ylmethylene)-hydrazine(Compound 3), 4-chloro-2-(3-hydroxypropyl)-6-morpholinopyrimidine (0.81g, 3.15 mmol) was synthesized, methylated with sodium hydride (0.48 g,6.30 mmol) for 10 min, and MeI (0.895 g, 6.30 mmol) for 5 h in 30 mL THFat 0° C. to give 4-chloro-2-(3-methoxypropyl)-6-morpholinopyrimidine ascolorless viscous oil (0.792 g, 3.03 mmol, 96%).

¹H NMR (300 MHz, CDCl₃), δ (ppm): 6.32 (s, 1H), 3.75-3.79 (m, 4H);3.61-3.64 (m, 4H); 3.44 (t, J=6.6 Hz, 2H); 3.34 (s, 3H); 2.78 (t, J=7.8Hz, 2H); and 2.00-2.09 (m, 2H).

MS (EST): m/z 262.1 (M+H).

Following the typical procedure,4-chloro-2-(3-methoxypropyl)-6-morpholinopyrimidine (0.783 g, 3.00 mmol)was treated with hydrazine and indole-3-carboxaldehyde sequentially toyield 0.89 g of Compound 5 (2.26 mmol, 75%).

¹H NMR (300 MHz, DMSO-d₆), δ (ppm): 11.46 (s, 1H); 10.64 (s, 1H); 8.26(s, 1H); 8.17-8.20 (m, 1H); 7.72 (d, J=2.4 Hz, 1H); 7.43 (dd, J=6.0 Hz,2.4 Hz, 1H); 7.15-7.21 (m, 2H); 6.16 (s, 1H), 3.70-3.73 (m, 4H);3.52-3.56 (m, 4H); 3.37 (t, J=6.9 Hz, 2H); 3.23 (s, 3H); 2.50-2.57 (m,2H), and 1.88-1.97 (m, 2H).

MS (ESI) m/z 395.2 (M+H).

EXAMPLE 6 Preparation of Compound 6:3-{4-[N′-(1H-indol-3-ylmethylene)-hydrazino]-6-morpholin-4-yl-pyrimidin-2-ylsulfanyl}-propan-1-ol

Compound 6 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, DMSO-d₆), δ (ppm): 11.48 (s, 1H); 10.68 (s, 1H); 8.26(s, 1H); 8.15-8.18 (m, 1H); 7.73 (d, J=2.1 Hz, 1H); 7.42-7.44 (m, 1H);7.16-7.20 (m, 2H); 6.04 (s, 1H), 4.53 (t, J=5.1 Hz, 1H); 3.65-3.71 (m,4H); 3.48-3.56 (m, 6H); 3.06 (t, J=7.2 Hz, 2H), and 1.76-1.85 (m, 2H).

MS (ESI): m/z 413.1 (M+H).

EXAMPLE 7 Preparation of Compound 7:3-{2-[N′-(1H-indol-3-ylmethylene)-hydrazino]-6-morpholin-4-yl-pyrimidin-4-ylsulfanyl}-propan-1-ol

Compound 7 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, DMSO-d₆), δ (ppm): 11.34 (s, 1H); 10.48 (s, 1H); 8.45(d, J=7.8 Hz, 1H); 8.25 (s, 1H); 7.64 (d, J=2.7 Hz, 1H); 7.40 (d, J=8.1Hz, 1H); 7.05-7.19 (m, 2H); 6.08 (s, 1H), 4.60 (t, J=5.1 Hz, 1H);3.50-3.68 (m, 10H); 3.20-3.30 (m, 2H); and 1.78-1.86 (m, 2H).

MS (ESI): m/z 413.1 (M+H).

EXAMPLE 8 Preparation of Compound 8:N-[2-(2,2-dimethyl-[1,3]dioxolan-4-ylmethoxy)-6-morpholin-4-yl-pyrimidin-4-yl]-N′-(1H-indol-3-ylmethylene)-hydrazine

Compound 8 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.38 (br s, 1H); 8.30 (dd, J=7.2, 1.8,1H), 8.02 (br s, 1H); 8.00 (s, 1H); 7.44-7.41 (m, 2H); 7.32-7.26 (m,2H); 6.14 (s, 1H); 4.51-4.42 (m, 2H);, 4.22-4.12 (m, 2H); 3.96-3.91 (m,1H); 3.84-3.79 (m, 4H); 3.70-3.64 (m, 4H); 1.47 (s, 3H); and 1.38(s,3H).

MS (ESI): m/z 453.2 (M+H).

EXAMPLE 9 Preparation of Compound 9:N-{2-[2-(3,4-dimethoxy-phenyl)-ethoxy]-6-morpholin-4-yl-pyrimidin-4-yl}-N′-(1H-indol-3-ylmethylene)-hydrazine

Compound 9 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.43 (bs, 1H); 8.30 (d, J=7.5 Hz 1H);8.2 (bs, 1H); 8.02 (d, J=2.7 Hz, 1H); 7.46-7.40 (m, 2H); 7.30-7.26 (m,2H); 6.82 (d, J=1 Hz, 3H); 4.45 (d, J=3.6 Hz, 1H); 4.45 (t, J=5.2 Hz,2H); 3.87 (d, J=3.9 Hz, 3H); 3.86 (d, J=3.9 Hz, 3H); 3.81 (s, 4H);3.67(s, 4H); and 3.04 (t, J=5.0 Hz, 2H).

MS (ESI): m/z 503.2 (M+H).

EXAMPLE 10 Preparation of Compound 10:N-(1H-indol-3-ylmethylene)-N′-[6-morpholin-4-yl-2-(2-pyridin-2-yl-ethoxy)-pyrimidin-4-yl]-hydrazine

Compound 10 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 9.3 (bs 1H); 8.66 (s, 1H); 8.55-8.53(m, 1H); 8.28-8.26 (m, 1H); 8.04 (s, 1H); 7.62-7.57 (m, 1H); 7.41-7.10(m, 6H); 6.08 (s, 1H); 4.64 (t, J=6.6 Hz, 2H); 3.76 (s, 4H); 3.62 (s,4H); and 3.26 (t, J=6.6 Hz, 2H).

MS (ESI): m/z 444.2 (M+H).

EXAMPLE 11 Preparation of Compound 11:N-(1H-indol-3-ylmethylene)-N′-[6-morpholin-4-yl-2-(3-pyridin-2-yl-propyl)-pyrimidin-4-yl]-hydrazine

Compound 11 was prepared in a similar manner as described in Example 1.

¹H NMR (300 MHz, DMSO-d₆), δ (ppm): 11.47 (s, 1H); 10.65 (s, 1H);8.50(d, J=4.5 Hz, 1H); 8.26 (s, 1H); 8.20-8.18 (m, 1H); 7.72-7.68 (m,2H); 7.45-7.42 (m, 1H); 7.29-7.18 (m, 4H); 6.17(s, 1H); 3.73 (s, 4H);3.5 (s, 4H); 2.79 (t, J=7.5 Hz, 2H); 2-58-2.51 (m, 2H); and 2.18-2.06(m, 2H).

MS (ESI): m/z 442.2 (M+H).

EXAMPLE 12 Preparation of Compound 12:N-(3-methyl-benzylidene)-N′-[6-morpholin-4-yl-2-(2-pyridin-2-yl-ethoxy)-pyrimidin-4-yl]-hydrazine

Compound 12 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, CDCl₃), Λ (ppm): 8.55-8.48 (m, 2H); 7.71 (s, 1H);7.65-7.55 (m, 1H); 7.49-7.42 (m, 2H); 7.30-7.15 (m, 4H); 6.08 (s, 1H);4.64 (t, J=6.6 Hz, 2H); 3.81-3.75 (m, 4H); 3.64-3.61 (m, 4H); 3.25 (t,J=7.0 Hz, 2H); and 2.38 (s, 3H).

MS (ESI): m/z 419.2 (M+H).

EXAMPLE 13 Preparation of Compound 13:N-(3-ethyl-benzylidene)-N′-[6-morpholin-4-yl-2-[2-pyridin-2-yl-ethoxy)-pyrimidin-4-yl]-hydrazine

Compound 13 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.58-8.50 (m, 1H); 8.43 (s, 1H); 7.95(s, 1H); 7.64-7.58 (m, 2H); 7.30-7.25 (m, 1H); 7.18-7.05 (m, 3H);6,07(s, 1H); 4.65 (t, J=6.9 Hz, 2H); 3.80-3.76 (m, 4H); 3.64-3.61(m,4H); 3.26(t, J=6.9 Hz, 2H); 2.40 (q, J=7.6 Hz, 2H); and 1.45 (t, J=7.6Hz, 3H).

MS (EST): m/z 433.3 (M+H).

EXAMPLE 14 Preparation of Compound 14:N-(3-methyl-benzylidene)-N′-{6-morpholin-4-yl-2-(3-pyridin-2-yl-propyl)-pyrimidin-4-yl]-hydrazine

Compound 14 was prepared in a similar manner as described in Example 1.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 9.6 (bs, 1H); 8.53 (d, J=4.5 Hz, 1H);7.76 (s, 1H); 7.56 (t, J=6 Hz, 1H); 7.49-7.47 (m, 2H); 7.28 (m, 1H);7.18-7.06 (m, 3H); 6.26 (s, 1H); 3.81-3.79 (m, 4H); 3.69-3.67 (m, 4H);2.89 (t, J=7.8 Hz, 2H); 2.71 (t, J=7.5 Hz, 2H); 2.39 (s, 3H); and 2.22(t, J=7.5 Hz, 2H).

MS (ESI): m/z 417.2 (M+H).

EXAMPLE 15 Preparation of Compound 15:N-[6-morpholin-4-yl-2-(2-pyridin-2-yl-ethoxy)-pyrimidin-4-yl]-N′-(1-m-tolyl-ethylidene)-hydrazine

Compound 15 was prepared in a similar manner as described in Example 2.¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.56 (bs, 1H), 7.66-7.46 (m, 4H),7.32-7.26 (m, 2H), 7.16-7.14 (m, 2H), 6.44(s, 1H), 4.69 (t, J=6.9 Hz,2H), 3.80-3.77 (m, 4H), 3.63-3.60 (m, 4H), 3.31 (t, J=6.9 Hz, 2H), 2.39(s, 3H).

MS (ESI): m/z 433.2 (M+H).

EXAMPLE 16 Preparation of Compound 16:N-[1-(1H-indol-3-yl)-ethylidene]-N′-6-morpholin-4-yl-2-(2-pyridin-2-yl-ethoxy)-pyrimidin-4-yl]-hydrazine

Compound 16 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 9.35 (bs, 1H); 8.54 (dd, J=0.9, 4.2Hz, 1H); 8.33 (d, J=7.5 Hz, 1H); 7.93 (s, 1H); 7.58 (t, J=7.2 Hz, 1H);7.36-7.33 (m, 2H); 7.27-7.120 (m, 4H); 6.49 (s, 1H); 4.6 8(t, J=7.2 Hz,2H); 3.76-3.73 (m, 4H); 3.60-3-57 (m, 4H); 3.50 (s, 3H); and 3.33-3.28(t, J=7.0 Hz, 2H).

MS (ESI): m/z 458,2 (M+H).

EXAMPLE 17 Preparation of Compound 17: 3-Methyl-benzaldehydeO-[6-morpholin-4-yl-2-(2-pyridin-2-yl-ethoxy)-pyrimidin-4-yl]-oxime

Compound 17 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.56-8.53 (m, 1H); 8.45 (s, 1H);7.62-7.50 (m, 3H); 7.38-7.26 (m, 3H); 7.18-7.10 (m, 1H); 6.17 (s, 1H);4.68 (t, J=6.9 Hz, 2H); 3.80-3.76 (m, 4H); 3.67-3.64 (m, 4H); 3.29 (t,J=6.9 Hz, 2H); and 2.41 (s, 3H).

MS (ESI): m/z 420.1 (M+H).

EXAMPLE 18 Preparation of Compound 18: 1H-indole-3-carbaldehydeO{6-morpholin-4-yl-2-(2-pyridin-2-yl-ethoxy)-pyrimidin-4-yl]-oxime

Compound 18 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, DMSO-d₆), δ (ppm): 11.82 (bs, 1H); 8.81 (s, 1H); 8.50(d, J=4.5 Hz, 1H); 8.04 (d, J=6.9 Hz, 1H); 7.93(s, 1H); 7.72 (t, J=6.9Hz, 1H); 7.49 (d, J=6.9 Hz, 1H); 7.33 (d, J=7.8 Hz, 1H); 7.30-7.18 (m,3H); 6.22 (s, 1H); 4.57 (t, J=6.3 Hz, 2H); 3.67 (s, 4H); 3.56 (s, 4H);and 3.15 (t, J=6.3 Hz, 2H).

MS (ESI): m/z 445.2 (M+H).

EXAMPLE 19 Preparation of Compound 19:N-(1H-indol-3-ylmethylene)-N-{6-morpholin-4-yl-2-[2-(pyridin-3-yloxy)-ethoxy]-pyrimidin-4-yl}-hydrazine

Compound 19 was prepared in a similar manner as described in Example 2.

¹H NMR: (300 MHz, CDCl₃), δ (ppm): 9.20 (br s, 1H); 8.30 (br s, 1H);8,29 (t, J=3.3 Hz, 1H); 8.18-8.12 (m, 2H); 7.44-7.41 (m, 2H); 7.26-7.18(m, 5H); 6.08 (s, 1H); 4,66 (t, J=4.8 Hz, 2H); 4.29 (t, J=5.0 Hz, 2H);3.80-3.76 (m, 4H); and 3.67-3.62 (m, 4H).

MS (ESI): m/z 460.2 (M+H).

EXAMPLE 20 Preparation of Compound 20:N-(3-methyl-benzylidene)-N′-{6-morpholin-4-yl-2-[2-(pyridin-3-yloxy)-ethoxy]-pyrimidin-4-yl}-hydrazine

Compound 20 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.55 (s, 1H); 8,34 (br s, 1H);8.30-8.23 (m, 1H); 7.78 (s, 1H); 7.50-7.47 (m, 2H); 7.32-7.24 (m, 1H);7.20-7.17 (m, 3H); 6.14 (s, 1H); 4.66 (t, J=5.0 Hz, 2H); 4.35 (t, J=4.8Hz, 2H); 3.83-3.80 (m, 4H); 3.68-3.65 (m, 4H); and 2.40(s, 3H).

MS (ESI): m/z 435.2 (M+H).

EXAMPLE 21 Preparation of Compound 21:Butyl-{4-[N′-(1H-indol-3-ylmethylene)-hydrazino]-6-morpholin-4-yl-pyrimidin-2-yl}-amine

Compound 21 was prepared in a similar manner as described in Example 2.

¹H NMR (300 MHz, CDCl₃), δ ppm: 8.41 (bs, 1H), 8.33-8.30 (m, 1H), 8.19(bs, 1H), 7.95 (s, 1H), 7.41-7.37 (m, 2H), 7.29-7.25 (m, 2H), 5.96(s,1H), 4.65 (t, J=4 Hz, 1H), 3.83-3.80 (m, 4H), 3.65-3.62 (m, 4H), 3.36(dd, J=6.3, 13.5 Hz, 2H), 1.60-1.55 (m, 2H), 1.35-1.33 (m, 4H),0.92-0.87 (m, 3H).

MS (ESI): m/z 408.2 (M+H).

EXAMPLE 22 Preparation of Compound 22:N-(3-Methyl-benzylidene)-N′-6-morpholin-4-yl-2-(pyridin-3-yloxy)-pyrimidin-4-yl]-hydrazine

To a solution of 3-hydroxypyridine (950 mg, 10 mmol) in anhydrous THF(50 mL) at 0° C. under the nitrogen protection was added NaH (60% inoil) (480 mg, 12 mmol). The suspension was stirred for 0.5 h at 0° C.,and 2,4,6-trichloropyrimidine (1.84 g, 10 mmol) was added. After themixture warmed to room temperature and stirred for 2 h, the reaction wasquenched by ice brine and extracted with EtOAc (300 mL). The organicphase was washed with brine, dried (Na₂SO₄), filtered, evaporated invacuo. The cure product was purified by flash chromatography on a columnof silica gel (EtOAc-Hexane, 1:7). The product (1.80 g, 7.4 mmol) inCH₂Cl₂ (150 mL) at 0° C. was added slowly morpholine (2.5 g, 28 mmol).The reaction mixture was stirred at 0° C. for 1 h and another 1 h atroom temperature. The mixture was washed with water. The organic phasewas dried (Na₂SO₄), filtered and evaporated in vacuo and presented threeisomers. The isomers was separated by flash chromatography on a columnof silica gel (EtOAc-Hexane, 1:7 and 1:3) to obtain4-[6-chloro-2-(pyridin-3-yloxy)-pyrimidin-4-yl]-morpholine (320 mg,14.7%).

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.51(d, 1H), J=2.7 Hz), 8.44(dd, 1H,J=1.5, J=3.3 Hz), 7.53-7.49(m,1H), 7.34-7.3 (m, 1H), 6.25 (s, 1H),3.71-3.67(m, 4H), 3.51-3.48(m, 4H).

MS (EST): m/z 293.1.

To a solution of4-[6-chloro-2-(pyridin-3-yloxy)-pyrimidin-4-yl]-morpholine (295 mg, 1mmol) in THF (10 mL) was added anhydrous hydrazine (0.320 ml, 10 mmol)under the nitrogen protection. The mixture was heated at 70° C. for 15min. After cooling to room temperature, the reaction mixture wasquenched by ice brine and extracted with EtOAc (100 mL). The organicphase was washed with brine (10 mL) and water (10 ml×2), dried (Na₂SO₄),filtered, evaporated, and purified by flash chromatography on a columnof silica gel (CH₂Cl₂ and CH₂Cl₂-MeOH, 95:5) and to give[6-morpholin-4-yl-2-(pyridin-3-yloxy)-pyrimidin-4-yl]-hydrazine (180 mg)in 62% yield. M/Z (M+1) 289.2

To a solution of[6-morpholin-4-yl-2-(pyridin-3-yloxy)-pyrimidin-4-yl]-hydrazine (180 mg)(145 mg, 0.5 mmol) and m-tolylaldehyde (72 mg, 0.6 mmol) in MeOH (10 mL)was added acetic acid (1 drop). The reaction mixture was stirred at roomtemperature for 12 h and white solid was precipitated. The resultingprecipitate was collected by filtration and washed with little amount ofmetanol and to give 125 mg of Compound 22 in 64% yield.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.71(s, 1H), 8.57(d, 1H, J=2.4 Hz),8.44(dd, 1H, J=1.5, 3.2 Hz), 7.78(s,1H), 7.56-7.52(m,1H), 7.46-7.43(m,2H), 7.34-7.26(m, 2H), 7.17(d, 1H, J=8.1 Hz), 6.17 (s, 1H), 3.76-3.73(m,4H), 3.57-3.54(m, 4H), 2.38(s, 3H).

MS (ESI): in/z 391.2.

EXAMPLE 23 Preparation of Compound 23:N-(3-Methylbenzlidene)-N′-(5-methyl-6-morpholin-4-yl-2-phenylpyrimidin-4-yl)hydrazine

Benzamidine hydrochloride (7.06 g, 0.045 mol) and dimethylmethylmalonate (6.0 g, 0.041 mol) were dissolved in methanol (100 mL).Sodium methoxide (21.5 mL, 0.099 mol, 25 wt % solution in methanol) wasadded and the solution was stirred at room temperature for 18 h. Thevolume of solvent was reduced to approximately 50 mL under reducedpressure, then poured onto ice water. This solution was neutralized withHOAc which produced a white precipitate. This precipitate was collectedand dried to produce a white solid (6.1 g, 74%).

¹H NMR (DMSO-d₆) δ (ppm)1.68 (s, 3H), 7.70-7.87 (m, 3H), 8.21 (d, J=8.4Hz).

MS (ESI): m/z 203.1 (M+H)⁺

5-Methyl-2-phenyl-pyrimidine-4,6-diol (3.3 g, 0.016 mol) and POCl₃ wereheated to 60 C for 3 hrs. The solution was allowed to cool to roomtemperature then poured onto ice. The resultant white precipitate wasfiltered and dried to produce the desired compound as a white solid (810mg, 21%). ¹H NMR (DMSO-d₆) δ (ppm) 2.40 (s, 3H), 7.51-7.56 (m, 3H), 8.23(d, 8.4 Hz).

MS (ESI): m/z 239.1 (M+H)⁺

4,6-Dichloro-5-methyl-2-phenylpyrimidine (2.5 g, 0.010 mol) andmorpholine (2.93 g, 0.031 mol) were dissolved in THF (50 mL) and heatedto reflux for 3 hrs. The solution was allowed to cool then EtOAc (100mL) and water (100 mL) were added. The EtOAc layer was washed with water(3×100 mL), dried over MgSO₄, filtered and solvent was removed underreduced pressure. The resultant solid was used without furtherpurification (2.66 g, 92%).

MS (EST): m/z 298.1 (M+H)⁺

4-(6-Chloro-5-methyl-2-phenylpyrimidin-4-yl)morpholine (439 mg, 1.51mmol) was dissolved in THF (50 mL). Hydrazine (0.25 mL, 7.96 mmol) wasadded and the solution was heated to reflux for 18 hrs. The reaction wasallowed to cool the solvent was removed under reduced pressure. EtOAc(100 mL) and water (100 mL) were added. The EtOAc layer was washed withwater (3×100 mL), dried over MgSO₄, filtered and solvent was removedunder reduced pressure to produce a white solid (374 mg). This solid wasredissolved in THF (50 mL) and m-tolualdehyde (157 mg, 1.31 mmol) wasadded. The solution was heated to reflux for 4 hrs then allowed to cool.Solvent was removed under reduced pressure then EtOAc (100 mL) and water(100 mL) were added. The EtOAc layer was washed with water (3×100 mL),dried over MgSO₄, filtered and solvent was removed under reducedpressure. The crude product was purified by silcagel columnchromatography, eluting with 25% EtOAc/hexane to produce the puredesired product as a yellow solid (313 mg, 53%). ¹H NMR (DMSO-d₆) δ(ppm) 2.26 (s, 3H), 2.36 (s, 3H), 3.35 (m, 4H), 3.75-3.78 (m, 4H), 7.20(d, J=6.9 Hz), 7.33 (t, J=6.9 Hz), 7.47-7.52 (m, 5H), 8.19 (s, 1H),8.35-8.38 (m, 2H), 10.60 (s, 1H).

MS (ESI): m/z 388.3 (M+H)⁺

EXAMPLE 24 Preparation of Compound 24:N-(3-methyl-benzylidene)-N′-(2-phenyl-6-thiomorpholin-4-yl-pyrimidin-4-yl)-hydrazine

Compound 24 was prepared in a similar manner as described in Example 23.

¹H-NMR (DMSO-d₆) δ 2.36 (s, 3H), 2.76 (s, 4H), 4.07 (s, 4H), 6.36 (s,1H), 7.19 (d, J=8.1 Hz), 7.32 J=8.1 Hz), 7.47-7.57 (m, 5H), 8.09 (s,1H), 8.30-8.31 (m, 1H), 11.02 (s, 1H).

MS (ESI): m/z 389.1.

EXAMPLE 25 Preparation of Compound 25:(2,3-Dimethyl-1H-indole-5-yl)-{6-morpholin-4-yl-2-[2-(pyridin-3-yloxy)-ethoxyl-pyrimidin-4-yl}-amine

To a solution of 2-(pyridin-3-yloxy)-ethanol (3.48 g, 25 mmol) in 40 mLof anhydrous THF at room temperature under the N₂, 2,4,6-trichloropyrimidine (4.56 g, 25 mmol) was added followed by portionwise additionof NaH (60% suspension in oil, 1.1 g, 27.5 mmol). After 30 min ofstirring reaction was quenched with water, water layer extracted withEtOAc, combined organic solutions washed with brine and dried overMgSO₄.

Purification using flash chromatography (silica;dichloromethane/acetone/methanol: 3/1/0.1) afforded mixture of4,6-dichloro-2- and2,6-dichloro-4-[2-(pyridin-3-yloxy)-ethoxy]-pyrimidines (3.72 g, 52%),(NMR ratio 1:1.2) as an oil.

To a solution of the above mixture (3.72 g, 13 mmol) in 20 mL of1,4-dioxane was added DIPEA (2.49 mL, 14.3 mmol), followed by2,3-dimethyl-5-amino-indole (2.08 g, 13 mmol) and a mixture was refluxedfor 1 hour. Solvent was removed under reduced pressure and reactionmixture was separated using column chromatography (silica;dichloromethane/acetone/methanol: 3/1/0.1) to afford{6-chloro-2-[2-(pyridin-3-yloxy)-ethoxy}-pyrimidin-4-yl}-amine (2.07 g,39%). An mixture of{4-chloro-6-[2-(pyridin-3-yloxy)-ethoxy]-pyrimidin-4-yl}-amine and{2-chloro-6-[2-(pyridin-3-yloxy)-ethoxy]-pyrimidin-4-yl}-amine (2.5 g,47%) was also obtained and used in another reaction.

A solution of{6-chloro-2-[2-(pyridin-3-yloxy)-ethoxy]-pyrimidin-4-yl}-amine (2.07 g,5.05 mmol) and morpholine (1.32 mL, 15.15 mmol) in 1,4-dioxane washeated at 110° C. for 24 hours. Solvent was removed under reducedpressure and reaction mixture was purified using flash chromatography(silica; dichloromethane/acetone/methanol: 3/1/0.1) to afford Compound25 (2 g, 86%) as a colorless solid.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.34 (br s, 1H), 8.23 (dd, 1H, J=3.6,2.1), 7.96 (brs, 1H), 7.34-7.21 (m, 4H), 6.98(dd, 1H, J=8.4, 1.8 Hz),6.60 (brs, 1H), 5.36 (s, 1H), 4.65 (t, 2H, J=5.1 Hz), 4.34 (t, 2H, J=5.1Hz), 3.66 (m, 4H), 3.42 (m, 4H), 2.37(s, 3H), and 2.20 (s, 3H).

MS (ESI): m/z 461.5 (M+H).

EXAMPLE 26 Preparation of Compound 26:(2,3-Dimethyl-1H-indole-5-yl)-{4-morpholin-4-yl-6-[2-(pyridin-3-yloxy)-ethoxyl-pyrimidin-2-yl}-amine

Reaction of a mixture of{4-chloro-6-[2-(pyridin-3-yloxy)-ethoxyl-pyrimidin-4-yl}-amine and{2-chloro-6-[2-(pyridin-3-yloxy)-ethoxy]-pyrimidin-4-yl}-amine (2.5 g,47%) and (2.5 g, 6.1 mmol) with morpholine was carried out as describedin Example 24.

Purification by flash chromatography and recrystallization fromether-pentane gave 0.3 g of Compound 26.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.36 (br s, 1H), 8.24 (m, 1H), 7.85(m, 1H), 7.70 (brs, 1H), 7.26-7.14 (m, 4H), 6.78 (brs, 1H), 5.42 (s,1H), 4.68 (t, 2H, J=5.1), 4.31 (t, 2H, J=5.1), 3.70 (m, 4H), 3.54 (m,4H), 2.35(s, 3H), and 2.18 (s, 3H).

MS (ESI): m/z 461.5 (M+H).

EXAMPLE 27 Preparation of Compound 27:3-{4-[N′-(3-Methyl-benzylidene)-hydrazino]-6-morpholin-4-yl-pyrimidin-2-yl}-propionicacid ethyl ester

Compound 27 was prepared in a similar manner as described in Example 1.

¹H NMR (300 MHz, CDCl₃), δ (ppm): 8.22 (s, 1H); 7.69(s, 1H); 8.07 (s,1H); 7.47 (m, 2H); 7.28 (t, J=7.5 Hz, 1H); 7.17 (d, J=7.5 Hz, 1H);6.23(s, 1H); 4.13 (q, J=7.2 Hz, 2H); 3.78-3.81 (m, 4H); 3.62-3.65 (m,4H); 2.98 (t, J=7.2 Hz, 2H); 2.77 (t, J=7.2 Hz, 2H); 2.39 (s, 3H); 1.24(t, J=7.2 Hz, 3H).

MS (ESI): m/z 398.2 (M+H).

EXAMPLE 28 In Vitro Assays

Reagents. Staphylococcus aureus Cowan I (SAC) was obtained fromCalbiochem (La Jolla, Calif.), and lipopolysaccharide (LPS, Serratiamarscencens) was obtained from Sigma (St. Louis, Mo.). Human and mouserecombinant IFNγ were purchased from Boehringer Mannheim (Mannheim,Germany) and Pharmingen (San Diego, Calif.), respectively.

Human In Vitro Assay. Human PBMC were isolated by centrifugation usingFicoll-Paque (Pharmacia Biotech, Uppsala, Sweden) and prepared in RPMImedium supplemented with 10% fetal calf serum (FCS), 100 U/mLpenicillin, and 100 μg/mL streptomycin. PBMC were plated in wells of a96-well plate at a concentration of 5×10⁵ cells/well, and primed byadding IFNγ (30 U/mL) for 22 h and stimulated by adding LPS (1 μg/mL),or by adding IFNγ (100 U/mL) and then stimulated by adding SAC (0.01%).A test pyrimidine compound was dissolved in DMSO, and added to wells ofthe 96-well plate. The final DMSO concentration was adjusted to 0.25% inall cultures, including the compound-free control. Human THP-1 cellswere plated in wells, primed by adding IFNγ (100 U/mL) for 22 h andstimulated by adding SAC (0.025%) in the presence of differentconcentrations of the pyrimidine compound. Cell-free supernatants weretaken 18 h later for measurement of cytokines. Cell viability wasassessed using the bioreduction of MTS. Cell survival was estimated bydetermining the ratio of the absorbance in compound-treated groupsversus compound-free control.

The supernatant was assayed for the amount of IL-12p40, IL-12p70, orIL-10 by using a sandwich ELISA with anti-human antibodies, i.e., aHuman IL-12 p40 ELISA kit from R&D Systems (Berkeley, Calif.), and aHuman IL-12 p70 or IL-10 ELISA kit from Endogen (Cambridge, Mass.).Assays were based on the manufacturer's instructions.

Murine In Vitro Assay. Balb/c mice (Taconic, Germantown, N.Y.) wereimmunized with Mycobacterium tuberculosis H37Ra (Difco, Detroit, Mich.).The splenocytes were harvested 5 days and prepared in RPMI mediumsupplemented with 10% FCS and antibiotics in a flat bottom 96-well platewith 1×10⁶ cells/well. The splenocytes were then stimulated with acombination of IFNγ (60 ng/mL) and SAC (0.025%) [or LPS (20 μg/mL)] inthe presence of a test compound. Cell-free supernatants were taken 24 hlater for the measurement of cytokines. The preparation of compound andthe assessment of cell viability were carried out as described above.Mouse IL-12 p70, IL-10, IL-1β, and TNFα were measured using ELISA kitsfrom Endogen, according to the manufacturer's instructions.

The biological activities of pyrimidine compounds were tested on humanPBMC or THP-1 cells. Many of the compounds have IC₅₀ values of 5 μM orless. Unexpectedly, some of the test compounds have IC₅₀ values as lowas 1 nM.

EXAMPLE 29 In Vivo Assays

Treatment of adjuvant arthritis in rats: Adjuvant arthritis (AA) wasinduced in female Lewis rats by the intracutaneous injection (base ofthe tail) of 0.1 mL of a 10 mg/mL bacterial suspension made from ground,heat-killed Mycobacterium tuberculosis H37Ra suspended in incompleteFreund's adjuvant. Rats were given a test compound orally once a day for12 days, starting the day following the induction. The development ofpolyarthritis was monitored daily by macroscopic inspection andassignment of an arthritis index to each animal, during the criticalperiod (days 10 to 25 post-immunization).

The intensity of polyarthritis was scored according to the followingscheme: (a) Grade each paw from 0 to 3 based on erythema, swelling, anddeformity of the joints: 0 for no erythema or swelling; 0.5 if swellingis detectable in at least one joint; 1 for mild swelling and erythema; 2for swelling and erythema of both tarsus and carpus; and 3 for ankylosisand bony deformity. Maximum score for all 4 paws was thus 12. (b) Gradefor other parts of the body: for each ear, 0.5 for redness and another0.5 if knots are present; 1 for connective tissue swelling (saddlenose); and 1 for the presence of knots or kinks in the tail. The highestpossible arthritic index was 16.

Experiments with the AA model were repeated four times. Oraladministration of pyrimidine compounds of this invention (e.g., Compound12) reproducibly reduced the arthritic score and delayed the developmentof polyarthritis in a dose-dependent manner. The arthritis score used inthis model was a reflection of the inflammatory state of the structuresmonitored and the results therefore show the ability of the testcompound to provide relief for this aspect of the pathology.

Treatment of Crohn is disease in dinitrobenzene sulfonic acid-inducedinflammatory bowel syndrome model rats: Wistar derived male or femalerats weighing 200±20 g and fasted for 24 hours were used. Distal colitiswas induced by intra-colonic instillation of 2,4-dinitrobenzene sulfonicacid (DNBS, 25 mg in 0.5 mL ethanol 30%) after which air (2 mL) wasgently injected through the cannula to ensure that the solution remainedin the colon. A test compound and/or vehicle was administered orally 24and 2 hours before DNBS instillation and then daily for 5 days. Onecontrol group was similarly treated with vehicle alone while the otheris treated with vehicle plus DNBS. The animals were sacrificed 24 hoursafter the final dose of test compound administration and each colon wasremoved and weighed. Colon-to-body weight ratio was then calculated foreach animal according to the formula: Colon (g)/BW×100. The “Net”increase in ratio of Vehicle-control+DNBS group relative toVehicle-control group was used as a base for comparison with testsubstance treated groups and expressed as “% Deduction.” Pyrimidinecompounds of this invention (e.g., Compound 12) reproducibly had about30% deduction. A 30% or more reduction in colon-to-body weight ratio,relative to the vehicle treated control group, was consideredsignificant.

Rats treated with test substance orally showed a marked reduction in theinflammatory response. These experiments were repeated three times andthe effects were reproducible.

Treatment of Crohn is disease in CD4⁺CD45Rb^(high) T cell-reconstitutedSCID colitis model mice: Spleen cells were prepared from normal femaleBALB/c mice. For cell purification, the following anti-mouse antibodieswere used to label non-CD4⁺ T cells: B220 (RA3-6B2), CD11b (M1/70), andCD8α (53-6.72). All antibodies were obtained from BioSource (Camarillo,Calif.). M450 anti-rat IgG-coated magnetic beads (Dynal, Oslo, Norway)were used to bind the antibodies and negative selection was accomplishedusing an MPC-1 magnetic concentrator. The enriched CD4⁺ cells were thenlabeled for cell sorting with FITC-conjugated CD45RB (16A, Pharmingen,San Diego, Calif.) and PE-conjugated CD4 (CT-CD4, Caltag, Burlingame,Calif.). CD4⁺ CD45RB^(high) cells were operationally defined as theupper 40% of CD45Rb-staining CD4⁺ cells and sorted under sterileconditions by flow cytometry. Harvested cells were resuspended at4×10⁶/mL in PBS and injected 100 μL intraperitoneally into female C.B-17SCID mice. Pyrimidine compounds of this invention (e.g., Compound 12)and/or vehicle was orally administered once a day, 5 days per week,starting the day following the transfer. The transplanted SCID mice wereweighed weekly and their clinical condition was monitored.

Colon tissue samples were fixed in 10% buffered formalin and embedded inparaffin. Sections (4 μm) collected from ascending, transverse, anddescending colon were cut and stained with hematoxylin and eosin. Theseverity of colitis was determined based on histological examination ofthe distal colon sections, whereby the extent of colonic inflammationwas graded on a scale of 0-3 in each of four criteria: crypt elongation,cell infiltration, depletion of goblet cells, and the number of cryptabscesses.

LP lymphocytes were isolated from freshly obtained colonic specimens.After removal of payer's patches, the colon was washed in Ca/Mg-freeHBSS, cut into 0.5 cm pieces and incubated twice in HBSS containing EDTA(0.75 mM), DTT (1 mM), and antibiotics (amphotericin 2.5 μg/mL,gentamicin 50 μg/mL from Sigma) at 37° C. for 15 min. Next, the tissuewas digested further in RPMI containing 0.5 mg/mL collagenase D, 0.01mg/mL DNase I (Boehringer Manheim), and antibiotics at 37° C. LP cellswere then layered on a 40-100% Percoll gradient (Pharmacia, Uppsala,Sweden), and lymphocyte-enriched populations were isolated from thecells at the 40-100% interface.

To measure cytokine production, 48-well plates were coated with 10 μg/mLmurine anti-CD3ε antibody (145-2C11) in carbonate buffer (PH 9.6)overnight at 4° C. 5×10⁵ LP cells were then cultured in 0.5 ml ofcomplete medium in precoated wells in the presence of 1 μg/mL solubleanti-CD28 antibody (37.51). Purified antibodies were obtained fromPharmingen. Culture supernatants were removed after 48 h and assayed forcytokine production. Murine IFNγ was measured using an ELISA kit fromEndogen (Cambridge, Mass.), according to the manufacturer'sinstructions.

Histological analysis showed that oral administration of pyrimidinecompounds of this invention (e.g., Compound 12) reduced colonicinflammation as compared to vehicle control. The suppressive effect wasdose-dependent with a substantial reduction at a dose of 10 mg/kg. Thecalculated colon-to-body weight ratio was consistent with thehistological score, showing attenuation by treatment with the testcompound. Furthermore, analysis of cytokines from LP cells in responseto anti-CD3 antibody and anti-CD28 antibody demonstrated that LP cellsfrom vehicle control produced an augmented level of IFNγ and treatmentwith test substance greatly diminished the production. These resultsclearly demonstrated the potential of the test substance in treatment ofinflammatory bowel disease represented by Crohn's disease.

EXAMPLE 30 Inhibition of IL-23

The compounds of the invention inhibit the expression of p40 that is asubunit of both IL-12 and IL-23. Therefore, inhibition of IL-23 inaddition to IL-12 is expected. In order to confirm the hypothesis, anassay was established to specifically detect IL-23 using polyclonalantibodies recognizing p19 (R&D Systems, MN), an IL-23 specific subunit.A 96-well plate was coated with the antibodies at and after washingincubated with the supernatants of human peripheral blood mononuclearcells (PBMC). The culture was stimulated with 1 μg/ml of liposaccharide(LPS) (FIG. 2) or 0.025% of S. aureus Cowan I (SAC) (FIG. 1) in thepresence of test compound after IFN-γ priming. The captured IL-23 wasthen detected by a biotinylated goat anti-human p40 antibody that bindsto p40 subunit of human IL-12 and IL-23 as a monomer or in the contextof the respective heterodimer (Part 840099 of product DY1240 from R&DSystems). The plate was developed by incubation with streptavidin-HRPand then substrate solution (R&D Systems Cat #DY999). Recombinant IL-23(R&D Systems) was added as standard in the assay. The estimateddetection range is from 0.1 to 10 ng/ml, and 1 ng/ml recombinant IL-12heterodimer (Cell Sciences, MA) and p40 monomer (R&D Systems) were underdetection limit. To compare with the inhibition of IL-23, thesupernatant was also analyzed for IL-12 heterodimer and total p40proteins using IL-12 specific ELISA kit (Cell Sciences) and p40 ELISAkit (R&D Systems) respectively. IL-23 was significantly induced inIFN-γ/SAC and IFN-γ/LPS-stimulated human PBMC, and was inhibited bycompound 12 in a dose-dependent manner. The inhibitory activity ofcompound 12 against IL-23 was comparable to that against p40 andslightly lower than that against IL-12.

EXAMPLE 31 Gene Expression of Peripheral Blood Mononuclear Cells AfterTreatment with a Compound of the Invention

Changes in gene expression patterns of peripheral blood mononuclearcells (PBMC) are studied using a gene chip microarrays (Affymetrix,Inc.). PBMC are stimulated with IFNγ plus SAC, then dosed with 0.1, 1.0,10, 100, or 1000 nM of a compound of the invention for 3 h. Control PBMCare stimulated with INFγ alone and IFNγ plus SAC. Changes in geneexpression patterns between the control samples and the samples dosedwith a compound of the invention are compared. In order to know thekinetics in the expression, PBMC with IFNγ/SAC are further studied atdifferent time points (20 min, 1.5 h, 3 h, 6 h, and 16 h) after theaddition of the stimulus. In addition, PBMC preparations can befractionated into T-cell enriched and monocyte-macrophage enrichedpopulations, in order to distinguish the effects of a compound of theinvention on these cell populations, following IFNγ/SAC stimulation.

Genes preferentially expressed in monocyte/macrophage cells includefirst and foremost, those encoding the p40 subunit of IL-12 and IL-23,as well as the p35 subunit of IL-12. The expression of EBI3 is inducedafter stimulation with IFNγ/SAC, and is expected to be dose-dependentlyinhibited by a compound of the invention because IL-27 is a heterodimerformed from subunits EBI3 and p28, and EBI3 shares 27% amino acidsequence homology with IL-12 p40 and p28 is a protein related to the p35subunit of IL-12.

Other Embodiments

All of the features disclosed in this specification may be combined inany combination. Each feature disclosed in this specification may bereplaced by an alternative feature serving the same, equivalent, orsimilar purpose. Thus, unless expressly stated otherwise, each featuredisclosed is only an example of a generic series of equivalent orsimilar features.

From the above description, one skilled in the art can easily ascertainthe essential characteristics of the present invention, and withoutdeparting from the spirit and scope thereof, can make various changesand modifications of the invention to adapt it to various usages andconditions. For example, compounds structurally analogous a pyrimidinecompound described in the specification also can be made, screened fortheir inhibiting IL-12 activities, and used to practice this invention.Thus, other embodiments are also within the claims.

1-44. (canceled)
 45. A method of treating a subject with an IL-10,and/or IL-12 related disorder selected from the group consisting ofadjuvant arthritis, inflammatory bowel disease, and Crohn's disease,said method comprising administering to the subject an effective amountof a compound of formula (I):

or a pharmaceutically acceptable salt thereof, wherein R₁ is

aryl, or heteroaryl; each of R₂ and R₄, independently, is R^(c),halogen, nitro, cyano, isothionitro, SR^(c), or OR^(c); or R₂ and R₄,taken together, is carbonyl. R₃ is R^(c), alkenyl, alkynyl, OR^(c),OC(O)R^(c), SO₂R^(c), S(O)R^(c), S(O₂)NR^(c)R^(d), SR^(c), NR^(c)R^(d),NR^(c)COR^(d), NR^(c)C(O)OR^(d), NR^(c)C(O)NR^(c)R^(d), NR^(c)SO₂R^(d),COR^(c), C(O)OR^(c), or C(O)NR^(c)R^(d); R₅ is H or alkyl; n is 0, 1, 2,3, 4, 5, or 6; X is O, S, S(O), S(O₂), or NR^(c); Y is a covalent bond,CH₂, C(O), C═N—R^(c), C═N—OR^(c), C═N—SR^(c), O, S, S(O), S(O₂), orNR^(c); Z is N or CH; one of U and V is N, and the other is CR^(c); andW is O, S, S(O), S(O₂), NR^(c), or NC(O)R^(c); in which each of R^(a)and R^(b), independently, is H, alkyl, aryl, heteroaryl; and each ofR^(c) and R^(d), independently, is H, alkyl, aryl, heteroaryl, cyclyl,heterocyclyl, or alkylcarbonyl.
 46. The method of claim 45, wherein R₁is


47. The method of claim 46, wherein U is N and V is CH.
 48. The methodof claim 46, wherein Z is N and W is O.
 49. The method of claim 46,wherein X is NR^(c).
 50. The method of claim 49, wherein R^(c) is H,methyl, ethyl, or acetyl.
 51. The method of claim 46, wherein Y is O orCH₂, and n is 0, 1, 2, 3, or
 4. 52. The method of claim 46, wherein oneof R^(a) and R^(b) is

wherein: B is NR^(i), O, or S; B′ is N or CR^(i); R^(g) is H, alkyl, oralkoxyl; R^(h) is halogen, NO₂, CN, alkyl, aryl, heteroaryl, OR^(c),OC(O)R^(c), SO₂R^(c), S(O)R^(B), S(O₂)NR^(c)R^(d), SR^(c), NR^(c)R^(d),NR^(c)COR^(d), NR^(c)C(O)OR^(d), NR^(c)O(O)NR^(c)R^(d), NR^(c)SO₂R^(d),COR^(c), C(O)OR^(c), or C(O)NR^(c)R^(d); R^(i) is H, alkyl, oralkylcarbonyl; p is 0, 1, or 2; and q is 0, 1, 2, 3, or
 4. 53. Themethod of claim 52, wherein one of R^(a) and R^(b) is

and the other of R^(a) and R^(b) is H or alkyl.
 54. The method of claim53, wherein R^(g) is H, methyl, ethyl, propyl, cyclopropyl, methoxy, orethoxy; R^(h) is F, Cl, CN, methyl, methoxy, ethoxy, OC(O)CH₃,OC(O)C₂H₅, C(O)OH, C(O)OC₂H₅, C(O)NH₂, NHC(O)CH₃, or S(O₂)NH₂; R^(i) isH, methyl, ethyl, or acetyl, and q is 0, 1, or
 2. 55. The method ofclaim 54, wherein R^(g) is methyl or methoxy; R^(i) is H; and q is 0.56. The method of claim 54, wherein U is N and V is CH.
 57. The methodof claim 56, wherein Z is N and W is O.
 58. The method of claim 57,wherein X is NR^(c); and R^(c) is H, methyl, ethyl, or acetyl.
 59. Themethod of claim 58, wherein Y is O or CH₂; and n is 0, 1, 2, 3, or 4.60. The method of claim 54, wherein Y is O or CH₂, and n is 0, 1, 2, 3,or
 4. 61. The method of claim 60, wherein R₃ is aryl or heteroaryl. 62.The method of claim 60, wherein R₃ is pyridinyl.
 63. The method of claim45, wherein R₁ is aryl or heteroaryl.
 64. The method of claim 45,wherein the compound is selected from the group consisting of:

or a pharmaceutically acceptable salt thereof.